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The aim of this study was to elucidate the effects of dietary pterostilbene supplementation on physicochemical changes and gel properties of myofibrillar protein (MP) in chicken when subjected to short-term frozen storage. The results showed that pterostilbene supplementation diminished the oxidation of MP compared to the control, as the carbonyl content was significantly reduced and the loss of sulfhydryl and free amino groups was slowed. Meanwhile, the surface hydrophobicity and insolubility of MP were significantly reduced. FT-IR and endogenous fluorescence spectroscopy analysis indicated that dietary pterostilbene inhibited the unfolding of protein structure and the transition of α-helix to β-sheet structure. The integrity of the protein structure contributed to the gel quality. The strength, whiteness and water-holding capacity (WHC) of MP gels were improved in the pterostilbene treatment group. In terms of microstructure, pterostilbene facilitated the formation of dense and homogeneous gel network structure. In summary, these findings suggest that pterostilbene could be used as a dietary supplement to maintain the structural stability of MP in postmortem chicken breast muscle, allowing for excellent gel functional properties.

The effects of ε-polylysine (ε-PL) at different concentrations (0.005 %, 0.010 %, 0.020 %, and 0.030 %) on the structure and gelling behavior of pork myofibrillar protein (MP) under oxidative stress were explored. The incorporation of ε-PL significantly restrained oxidation-induced sulfhydryl and solubility losses (up to 9.72 % and 41.9 %, respectively) as well as protein crosslinking and aggregation. Compared with the oxidized control, ε-PL at low concentrations (0.005 % - 0.020 %) promoted further unfolding and destabilization of MP, while 0.030 % ε-PL led to refolding of MP and enhanced its thermal stability. The ε-PL-induced physicochemical changes favored the formation of a finer and more homogeneous three-dimensional network structure, therefore obviously enhancing the strength and water-holding capacity (WHC) of thermally induced oxidized MP gels, with the ε-PL at 0.020 % showed the greatest enhancement. This work revealed for the first time that ε-PL can significantly ameliorate the oxidation stability and gel-forming ability of meat proteins.

FiberSim is a flexible open-source model of myofilament-level contraction. The code uses a spatially explicit technique, meaning that it tracks the position and status of each contractile molecule within the lattice framework. This allows the model to simulate some of the mechanical effects modulated by myosin-binding protein C, as well as the dose dependence of myotropes and the effects of varying isoform expression levels. This paper provides a short introduction to FiberSim and presents simulations of tension-pCa curves with and without regulation of thick and thin filament activation by myosin-binding protein C. A myotrope dose-dependent response as well as slack/re-stretch maneuvers to assess rates of tension recovery are also presented. The software was designed to be flexible (the user can define their own model and/or protocol) and computationally efficient (simulations can be performed on a regular laptop). We hope that other investigators will use FiberSim to explore myofilament level mechanisms and to accelerate research focusing on the contractile properties of sarcomeres.

In this study, we investigated the interaction between myofibrillar proteins (MPs) and selected alcohols (1-pentanol, 1-hexanol, and 1-heptanol). Only 1-heptanol exhibited the binding ability to MPs, and the binding ability significantly increased with increasing protein concentration (p 0.05). In addition, both static and dynamic quenching occurred during the interaction, with a red shift of the maximum absorption peak in the synchronous fluorescence spectra indicating a change in the microenvironment of the MPs. The results of circular dichroism measurements suggested that the interaction between MPs and 1-heptanol altered the secondary structure of the MPs. Furthermore, thermodynamic analysis showed that hydrogen bonding and van der Waals forces dominated the interaction between MPs and 1-heptanol, which was confirmed by the results of molecular docking/dynamics simulations. This study provides an in-depth understanding of the interaction between MPs and alcohols, which can help to improve the flavor control in meat.

Myofibrillar proteins (MPs) have not been fully used for a long time due to its poor solubility in low ionic strength solutions. The study explored the effect of high pressure homogenization (HPH) cycles under two pressures on the solubility of MPs. The MPs solubility increased with HPH cycles (p 0.05), the results of turbidity, appearance, droplet size indicated that the increase of solubility was due to MPs depolymerization, excessive HPH cycles (25k psi for 11 cycles) would lead to protein re-aggregation but does not affect solubility (p0.05). SDS-PAGE suggested that myosin formed soluble polymers with different molecular weights through disulfide bonds during HPH cycles, the polymer consisted of myosin subunits of different molecular weights. Endogenous fluorescence spectra, intermolecular chemical forces, isoelectric point analysis and free amino acids (FAAs) indicated that the dissolution of polymers in low ionic strength media was dominated by polar environment and intermolecular steric hindrance, but not to FAAs.

One of the complexities of understanding the pathology of familial forms of cardiac diseases is the level of mutation incorporation in sarcomeres. Computational models of the sarcomere that are spatially explicit offer an approach to study aspects of mutational incorporation into myofilaments that are more challenging to get experimentally. We studied two well characterized mutations of cardiac TnC, L48Q and I61Q, that decrease or increase the release rate of Ca(2+) from cTnC, k(−Ca), resulting in HCM and DCM respectively [1]. Expression of these mutations in transgenic mice was used to provide experimental data which showed incorporation of 30 and 50% (respectively) into sarcomeres. Here we demonstrate that fixed length twitch contractions of trabeculae from mice containing mutant differ from WT; L48Q trabeculae have slower relaxation while I61Q trabeculae have markedly reduced peak tension. Using our multiscale modelling approach [2] we were able to describe the tension transients of WT mouse myocardium. Tension transients for the mutant cTnCs were simulated with changes in k(-Ca), measured experimentally for each cTnC mutant in whole troponin complex, a change in the affinity of cTnC for cTnI, and a reduction in the number of detached crossbridges available for binding. A major advantage of the multiscale explicit 3-D model is that it predicts the effects of variable mutation incorporation, and the effects of variations in mutation distribution within thin filaments in sarcomeres. Such effects are currently impossible to explore experimentally. We explored random and clustered distributions of mutant cTnCs in thin filaments, as well as distributions of individual thin filaments with only WT or mutant cTnCs present. The effects of variable amounts of incorporation and non-random distribution of mutant cTnCs are more marked for I61Q than L48Q cTnC. We conclude that this approach can be effective for study on mutations in multiple proteins of the sarcomere. SUMMARY: A challenge in experimental studies of diseases is accounting for the effect of variable mutation incorporation into myofilaments. Here we use a spatially explicit computational approach, informed by experimental data from transgenic mice expressing one of two mutations in cardiac Troponin C that increase or decrease calcium sensitivity. We demonstrate that the model can accurately describe twitch contractions for the data and go on to explore the effect of variable mutant incorporation and localization on simulated cardiac muscle twitches.

The cardiac thin filament is regulated in a Ca2+-dependent manner through conformational changes of troponin and tropomyosin (Tm). It has been generally understood that under conditions of low Ca2+ the inhibitory peptide domain (IP) of troponin I (TnI) binds to actin and holds Tm over the myosin binding sites on actin to prevent crossbridge formation. More recently, evidence that the C-terminal mobile domain (MD) of TnI also binds actin has made for a more complex scenario. This study uses a computational model to investigate the consequences of assuming that TnI regulates Tm movement via two actin-binding domains rather than one. First, a 16-state model of the cardiac thin filament regulatory unit was created with TnI-IP as the sole regulatory domain. Expansion of this to include TnI-MD formed a 24-state model. Comparison of these models showed that assumption of a second actin-binding site allows the individual domains to have a lower affinity for actin than would be required for IP acting alone. Indeed, setting actin affinities of the IP and MD to 25% of that assumed for the IP in the single-site model was sufficient to achieve precisely the same degree of Ca2+ regulation. We also tested the 24-state model's ability to represent steady-state experimental data in the case of disruption of either the IP or MD. We were able to capture qualitative changes in several properties that matched what was seen in the experimental data. Lastly, simulations were run to examine the effect of disruption of the IP or MD on twitch dynamics. Our results suggest that both domains are required to keep diastolic cross-bridge activity to a minimum and accelerate myofilament relaxation. Overall, our analyses support a paradigm in which two domains of TnI bind with moderate affinity to actin, working in tandem to complete Ca2+-dependent regulation of the thin filament.

Summary Disease modeling and pharmaceutical testing using cardiomyocytes derived from induced pluripotent stem cells (iPSC-CMs) requires accurate assessment of contractile function. Micropatterning iPSC-CMs on elastic substrates controls cell shape and alignment to enable contractile studies, but determinants of intrinsic variability in this system have been incompletely characterized. The objective of this study was to determine the impact of myofibrillar structure on contractile function in iPSC-CMs. Automated analysis of micropatterned iPSC-CMs labeled with a cell-permeant F-actin dye revealed that myofibrillar abundance is widely variable among iPSC-CMs and strongly correlates with contractile function. This variability is not reduced by subcloning from single iPSCs and is independent of the iPSC-CM purification method. Controlling for myofibrillar structure reduces false-positive findings related to batch effect and improves sensitivity for pharmacologic testing and disease modeling. This analysis provides compelling evidence that myofibrillar structure should be assessed concurrently in studies investigating contractile function in iPSC-CMs., Highlights • iPSC-cardiomyocytes (iPSC-CMs) exhibit marked variability in contractile function • Myofibrillar structure and abundance correlates with iPSC-CM contractile function • Myofibrillar variability is not diminished by single-cell subcloning • Controlling for myofibrillar structure is important for iPSC-CM contractile studies, Contractile function is a critical phenotype of iPSC-derived cardiomyocytes (iPSC-CMs), but variability across cells limits application for disease modeling and pharmacologic testing. Using live-cell imaging of myofibrils and traction forces, Helms and colleagues demonstrate that contractile function is dependent on variable myofibrillar structure. Concurrent assessment of myofibrillar structure is therefore important for iPSC-CM contractile studies.

In this study, the microbiological, physicochemical, and flavor changes of turbot (Scophthalmus maximus) coated with a composite active coating of locust bean gum (LBG) and sodium alginate (SA) supplemented with daphnetin emulsions (0.16, 0.32, 0.64 mg·mL−1) were determined during 18 days of refrigerated storage (4 ± 1 °C). Results showed that LBG-SA coatings containing 0.32 mg·mL−1 daphnetin emulsions could significantly lower the total viable count (TVC), psychrophiles, Pseudomonas spp. and H2S-producing bacteria counts, and inhibit the productions of off-flavor compounds including the total volatile basic nitrogen (TVB-N), trimethylamine (TMA) and ATP-related compounds. 32 volatile compounds were identified by solid phase microextraction combined with gas chromatography-mass spectrometer method (SPME-GC/MS) during refrigerated storage and the treated turbot samples significantly lowered the relative content of fishy flavor compounds. Further, the LBG-SA coatings containing daphnetin could also delay the myofibril degradation of the turbot samples. These results indicated that the LBG-SA coatings with 0.32 mg·mL−1 daphnetin were a potential alternative way to improve the quality of turbot during refrigerated storage.

To demonstrate the roles of bacteria in fish protein degradation and related quality changes, three major grass carp spoilage bacteria were individually inoculated into grass carp flesh, and their effects on protein molecules, muscle structures, and quality indices were evaluated through peptidomics, optical microscopy and transmission electron microscope, and texture and water distribution analyses, respectively. Results showed that Pseudomonas putida degraded obscurin, nebulin, and titin, caused disarrangement of myofilaments and fragmented myofibers, and induced great loss of free water in muscle. Shewanella putrefaciens was active in hydrolyzing collagen and degraded both thick and thin filament proteins. Mutual separation of myofibers and severe texture softening were also observed in S. putrefaciens-inoculated samples. Aeromonas rivipollensis degraded myosin heavy chain and some thin filament proteins but less affected muscle microstructure and quality indices. Therefore, this study revealed the mechanisms of bacteria-induced grass carp protein degradation and provided guidance for developing quality control strategies.

The neuromuscular junction (NMJ) connects the motor neuron with myofibers allowing muscle contraction. Both aging and increased activity result in NMJ remodeling. Here, the effects of exercise were examined in young and aged soleus muscles. Using immunofluorescent staining procedures, cellular and active zone components of the NMJ were quantified following a treadmill running program. Immunofluorescence was employed to determine myofiber profiles (size and type). Two-way analysis of variance procedures with main effects of age and treatment showed that when analyzing NMJs at the cellular level, significant (p ≤ 0.05) effects were identified for age, but not treatment. However, when examining subcellular active zones, effects for exercise, but not for age, were detected. Myofiber cross-sectional area showed that aging elicited atrophy and that among younger muscles endurance exercise training yielded decrements in myofiber size. Conversely, among aged muscles training elicited whole muscle and myofiber trends (p < 0.10) toward hypertrophy. Thus, different components of the neuromuscular system harbor unique sensitivities to various stimuli enabling proper adaptations to attain optimal function under differing conditions.

In this study, different ultrasound frequencies, including 20 kHz, 23 kHz, and 20/23 kHz, were applied for the treatment of myofibrillar protein (MP) with different protein concentrations (20, 40, and 60 mg/mL). The structures and emulsifying properties of the treated and untreated MPs were compared. The MP structural changes were confirmed based on the active sulfhydryl content (ASC), surface hydrophobicity (SUH), particle size, and turbidity, and the degree of change decreased in the order of 20/23 kHz > 20 kHz ≈ 23 kHz > control (CON). Moreover, the MP emulsifying properties, determined based on the emulsifying activity index (EAI), emulsion stability index (ESI), and creaming stability, were in accordance with the structural changes. Notably, frequency superposition provided more external force through the resonance of cavitation bubbles to induce stronger stress response of myosin in MP, thereby promoting the change of protein structure and emulsifying properties. Results implied that MPs treated with different ultrasound frequencies, especially the superposition of two frequencies, have the potential for wide applications in emulsified-type meat products.

The sarcomere is the functional unit of cardiac muscle, essential for normal heart function. To date, it has not been possible to study, in real time, thin filament-based activation dynamics in live cardiac muscle. We report here results from a cardiac troponin C (TnC) FRET-based biosensor integrated into the cardiac sarcomere via stoichiometric replacement of endogenous TnC. The TnC biosensor provides, for the first time, evidence of multiple thin filament activating ligands, including troponin I interfacing with TnC and cycling myosin, during a cardiac twitch. Results show that the TnC FRET biosensor transient significantly precedes that of peak twitch force. Using small molecules and genetic modifiers known to alter sarcomere activation, independently of the intracellular Ca(2+) transient, the data show that the TnC biosensor detects significant effects of the troponin I switch domain as a sarcomere-activating ligand. Interestingly, the TnC biosensor also detected the effects of load-dependent altered myosin cycling, as shown by a significant delay in TnC biosensor transient inactivation during the isometric twitch. In addition, the TnC biosensor detected the effects of myosin as an activating ligand during the twitch by using a small molecule that directly alters cross-bridge cycling, independently of the intracellular Ca(2+) transient. Collectively, these results aid in illuminating the basis of cardiac muscle contractile activation with implications for gene, protein, and small molecule-based strategies designed to target the sarcomere in regulating beat-to-beat heart performance in health and disease.

Background Congenital heart disease is one of the most common cardiac malformation-related diseases worldwide. Some causative genes have been identified but can explain only a small proportion of all cases; therefore, the discovery of novel susceptibility genes and/or modifier genes for abnormal cardiac development remains a major challenge. Methods We used a single nucleotide polymorphism (SNP) array, and next-generation sequencing (NGS) was conducted to screen and quickly identify candidate genes. KIAA0196 knockout zebrafish and mice were generated by CRISPR/Cas9 to detect whether or how KIAA0196 deficiency would influence cardiac development. Results Homozygous, but not heterozygous, zebrafish and mice showed early embryonic lethality. At the embryonic stage, microscopic examination and dissection revealed pericardial edema and ventricle enlargement in homozygous zebrafish and obviously delayed cardiac development in heterozygous mice, while echocardiography and tissue staining showed that significantly decreased cardiac function, ventricle enlargement, myofibril loss, and significantly reduced trabecular muscle density were observed in adult heterozygous zebrafish and mice. Most importantly, immunostaining and electron microscopy showed that there was a significant increase in sarcomere structural disorganization and myofibril structural integrity loss in KIAA0196 mutants. Furthermore, substantial downregulation in other sarcomeric genes and proteins was detected and verified in a mouse model via transcriptome and proteomics analyses; these changes especially affected the myosin heavy or light chain (MYH or MYL) family genes. Conclusion We identified KIAA0196 for the first time as a susceptibility gene for abnormal cardiac development. KIAA0196 deficiency may cause abnormal heart development by influencing the structural integrity of myofibrils.

It has been accepted that the force produced by a skeletal muscle myofibril depends on its cross-sectional area but not on the number of active sarcomeres because they are arranged in series. However, a previous study performed by our group showed that blocking actomyosin interactions within an activated myofibril and depleting the thick filaments in one sarcomere unexpectedly reduced force production. In this study, we examined in detail how consecutive depletion of thick filaments in individual sarcomeres within a myofibril affects force production. Myofibrils isolated from rabbit psoas were activated and relaxed using a perfusion system. An extra microperfusion needle filled with a high-ionic strength solution was used to erase thick filaments in individual sarcomeres in real time before myofibril activation. The isometric forces were measured upon activation. The force produced by myofibrils with intact sarcomeres was significantly higher than the force produced by myofibrils with one or more sarcomeres lacking thick filaments (p < 0.0001) irrespective of the number of contractions imposed on the myofibrils and their initial sarcomere length. Our results suggest that the myofibril force is affected by intersarcomere dynamics and the number of active sarcomeres in series.

The troponin complex regulates the Ca(2+) activation of myofilaments during striated muscle contraction and relaxation. Troponin genes emerged 500–700 million years ago during early animal evolution. Troponin T (TnT) is the thin-filament–anchoring subunit of troponin. Vertebrate and invertebrate TnTs have conserved core structures, reflecting conserved functions in regulating muscle contraction, and they also contain significantly diverged structures, reflecting muscle type- and species-specific adaptations. TnT in insects contains a highly-diverged structure consisting of a long glutamic acid–rich C-terminal extension of ∼70 residues with unknown function. We found here that C-terminally truncated Drosophila TnT (TpnT–CD70) retains binding of tropomyosin, troponin I, and troponin C, indicating a preserved core structure of TnT. However, the mutant TpnT(CD70) gene residing on the X chromosome resulted in lethality in male flies. We demonstrate that this X-linked mutation produces dominant-negative phenotypes, including decreased flying and climbing abilities, in heterozygous female flies. Immunoblot quantification with a TpnT-specific mAb indicated expression of TpnT–CD70 in vivo and normal stoichiometry of total TnT in myofilaments of heterozygous female flies. Light and EM examinations revealed primarily normal sarcomere structures in female heterozygous animals, whereas Z-band streaming could be observed in the jump muscle of these flies. Although TpnT–CD70-expressing flies exhibited lower resistance to cardiac stress, their hearts were significantly more tolerant to Ca(2+) overloading induced by high-frequency electrical pacing. Our findings suggest that the Glu-rich long C-terminal extension of insect TnT functions as a myofilament Ca(2+) buffer/reservoir and is potentially critical to the high-frequency asynchronous contraction of flight muscles.

Objective Cardiac troponin I (cTnI) is an essential physiological and pathological regulator of cardiac relaxation. Significant to this regulation, the post-translational modification of cTnI through phosphorylation functions as a key mechanism to accelerate myofibril relaxation. Similar to phosphorylation, post-translational modification by acetylation alters amino acid charge and protein function. Recent studies have demonstrated that the acetylation of cardiac myofibril proteins accelerates relaxation and that cTnI is acetylated in the heart. These findings highlight the potential significance of myofilament acetylation; however, it is not known if site-specific acetylation of cTnI can lead to changes in myofilament, myofibril, and/or cellular mechanics. The objective of this study was to determine the effects of mimicking acetylation at a single site of cTnI (lysine-132; K132) on myofilament, myofibril, and cellular mechanics and elucidate its influence on molecular function. Methods To determine if pseudo-acetylation of cTnI at 132 modulates thin filament regulation of the acto-myosin interaction, we reconstituted thin filaments containing WT or K132Q (to mimic acetylation) cTnI and assessed in vitro motility. To test if mimicking acetylation at K132 alters cellular relaxation, adult rat ventricular cardiomyocytes were infected with adenoviral constructs expressing either cTnI K132Q or K132 replaced with arginine (K132R; to prevent acetylation) and cell shortening and isolated myofibril mechanics were measured. Finally, to confirm that changes in cell shortening and myofibril mechanics were directly due to pseudo-acetylation of cTnI at K132, we exchanged troponin containing WT or K132Q cTnI into isolated myofibrils and measured myofibril mechanical properties. Results Reconstituted thin filaments containing K132Q cTnI exhibited decreased calcium sensitivity compared to thin filaments reconstituted with WT cTnI. Cardiomyocytes expressing K132Q cTnI had faster relengthening and myofibrils isolated from these cells had faster relaxation along with decreased calcium sensitivity compared to cardiomyocytes expressing WT or K132R cTnI. Myofibrils exchanged with K132Q cTnI ex vivo demonstrated faster relaxation and decreased calcium sensitivity. Conclusions Our results indicate for the first time that mimicking acetylation of a specific cTnI lysine accelerates myofilament, myofibril, and myocyte relaxation. This work underscores the importance of understanding how acetylation of specific sarcomeric proteins affects cardiac homeostasis and disease and suggests that modulation of myofilament lysine acetylation may represent a novel therapeutic target to alter cardiac relaxation.

Succinylation is a post-translational modification of protein lysine residues with succinyl groups derived from succinyl CoA. Succinylation is considered a significant post-translational modification with the potential to impact protein function which is highly conserved across numerous species. The role of succinylation in the heart, especially in heart failure and myofibril mechanics, remains largely unexplored. Mechanical parameters were measured in myofibrils isolated from failing hearts of ischemic cardiomyopathy patients and non-failing donor controls. We employed mass spectrometry to quantify differential protein expression in myofibrils from failing ischemic cardiomyopathy hearts compared to non-failing hearts. In addition, we combined peptide enrichment by immunoprecipitation with liquid chromatography tandem mass spectrometry to quantitatively analyze succinylated lysine residues in these myofibrils. Several key parameters of sarcomeric mechanical interactions were altered in myofibrils isolated from failing ischemic cardiomyopathy hearts, including lower resting tension and a faster rate of activation. Of the 100 differentially expressed proteins, 46 showed increased expression in ischemic heart failure, while 54 demonstrated decreased expression in ischemic heart failure. Our quantitative succinylome analysis identified a total of 572 unique succinylated lysine sites located on 181 proteins, with 307 significantly changed succinylation events. We found that 297 succinyl-Lys demonstrated decreased succinylation on 104 proteins, while 10 residues demonstrated increased succinylation on 4 proteins. Investigating succinyl CoA generation, enzyme activity assays demonstrated that α-ketoglutarate dehydrogenase and succinate dehydrogenase activities were significantly decreased in ischemic heart failure. An activity assay for succinyl CoA synthetase demonstrated a significant increase in ischemic heart failure. Taken together, our findings support the hypothesis that succinyl CoA production is decreased and succinyl CoA turnover is increased in ischemic heart failure, potentially resulting in an overall decrease in the mitochondrial succinyl CoA pool, which may contribute to decreased myofibril protein succinylation in heart failure.

Aberrant regulation of myocardial force production represents an early biomechanical defect associated with sarcomeric cardiomyopathies, but the molecular mechanisms remain poorly defined. Here, we evaluated the pathogenicity of a previously unreported sarcomeric gene variant identified in a pediatric patient with sporadic dilated cardiomyopathy, and we determined a molecular mechanism. Trio whole-exome sequencing revealed a de novo missense variant in TNNC1 that encodes a p.I4M substitution in the N-terminal helix of cardiac troponin C (cTnC). Reconstitution of this human cTnC variant into permeabilized porcine cardiac muscle preparations significantly decreases the magnitude and rate of isometric force generation at physiological Ca(2+)-activation levels. Computational modeling suggests that this inhibitory effect can be explained by a decrease in the rates of cross-bridge attachment and detachment. For the first time, we show that cardiac troponin T (cTnT), in part through its intrinsically disordered C terminus, directly binds to WT cTnC, and we find that this cardiomyopathic variant displays tighter binding to cTnT. Steady-state fluorescence and NMR spectroscopy studies suggest that this variant propagates perturbations in cTnC structural dynamics to distal regions of the molecule. We propose that the intrinsically disordered C terminus of cTnT directly interacts with the regulatory N-domain of cTnC to allosterically modulate Ca(2+) activation of force, perhaps by controlling the troponin I switching mechanism of striated muscle contraction. Alterations in cTnC–cTnT binding may compromise contractile performance and trigger pathological remodeling of the myocardium.

Cell fusion is essential for the development of multicellular organisms, and plays a key role in the formation of various cell types and tissues. Recent findings have highlighted the varied protein machinery that drives plasma-membrane merger in different systems, which is characterized by diverse structural and functional elements. We highlight the discovery and activities of several key sets of fusion proteins that together offer an evolving perspective on cell membrane fusion. We also emphasize recent discoveries in vertebrate myoblast fusion in skeletal muscle, which is composed of numerous multinucleated myofibers formed by the fusion of progenitor cells during development.

Cardiac myosin–binding protein C (cMyBP-C) is a thick filament-associated protein that influences actin-myosin interactions. cMyBP-C alters myofilament structure and contractile properties in a protein kinase A (PKA) phosphorylation-dependent manner. To determine the effects of cMyBP-C and its phosphorylation on the microsecond rotational dynamics of actin filaments, we attached a phosphorescent probe to F-actin at Cys-374 and performed transient phosphorescence anisotropy (TPA) experiments. Binding of cMyBP-C N-terminal domains (C0–C2) to labeled F-actin reduced rotational flexibility by 20–25°, indicated by increased final anisotropy of the TPA decay. The effects of C0–C2 on actin TPA were highly cooperative (n = ∼8), suggesting that the cMyBP-C N terminus impacts the rotational dynamics of actin spanning seven monomers (i.e. the length of tropomyosin). PKA-mediated phosphorylation of C0–C2 eliminated the cooperative effects on actin flexibility and modestly increased actin rotational rates. Effects of Ser to Asp phosphomimetic substitutions in the M-domain of C0–C2 on actin dynamics only partially recapitulated the phosphorylation effects. C0–C1 (lacking M-domain/C2) similarly exhibited reduced cooperativity, but not as reduced as by phosphorylated C0–C2. These results suggest an important regulatory role of the M-domain in cMyBP-C effects on actin structural dynamics. In contrast, phosphomimetic substitution of the glycogen synthase kinase (GSK3β) site in the Pro/Ala-rich linker of C0–C2 did not significantly affect the TPA results. We conclude that cMyBP-C binding and PKA-mediated phosphorylation can modulate actin dynamics. We propose that these N-terminal cMyBP-C–induced changes in actin dynamics help explain the functional effects of cMyBP-C phosphorylation on actin-myosin interactions.

The present work investigated effects of high-pressure homogenization (HPH) pressure (0, 40, 80 and 120 MPa) on physicochemical, functional and rheological properties of clam myofibrillar protein (CMP). Results showed that HPH changed the CMP secondary and tertiary structures. Absolute zeta potential and protein solubility increased but particle size and turbidity of CMP decreased after HPH treatment. Both of emulsifying properties and foaming properties were significantly improved. The shear stress, apparent viscosity and the viscosity coefficients reduced, but flow index increased. Application of HPH improved the physicochemical, functional and rheological properties of CMP, and 120 MPa was the optimal pressure for modification.

To compare the differences between direct protein oxidation (PO) and lipid-derived PO, the myofibrillar protein (MP) of obscure pufferfish was oxidatively modified by the hydroxyl radical oxidizing system (HOS) and the lipid-oxidizing system (LOS). The degree of oxidation, structural characteristics, and oxidation sites in MP were assessed. The results showed there was no significant thiol loss in LOS, compared with a 77.64% loss observed in case of the HOS. The secondary structure of MP was more vulnerable to HOS, but the tertiary structure was more susceptible to LOS. The cross-linking was largely attributed to the reversible disulfide links in HOS and the irreversible covalent linkages in LOS. Six amino acids and 10 specific oxidant products were identified in HOS. Only three amino acids and three specific oxidant products were identified in LOS. These findings may help deepen the understanding regarding the mechanism underlying PO in protein- and lipid-rich food materials.

This study aimed to facilitate the understanding on the origin of thawing drip under different freezing rate. Eventually we observed significantly greater thaw loss produced by slow freezing (8.58%) as compared to fast freezing (6.41%) after 24 h of thawing. Back to the freezing, ice crystallization induced decline in pH and the cold denaturation of myofibrillar protein. However, independent of freezing rate, we noticed protein renaturation with pH restoring during thawing, evidenced by the decreasing surface hydrophobicity, increasing solubility and thermal stability, and gradually stabilized secondary structure. Meanwhile, the water-holding of myofibrils increased with thawing process along with the rising water mobility. Under fast freezing, the results indicated less extensive protein cold denaturation and lower water mobility during thawing. Besides, we proposed that the microenvironment of lower ionic strength in fast freezing should benefit the protein renaturation and water re-absorption, ultimately contributed to lower thaw loss.

This study aimed to evaluate the effect of different aging times (1, 6 and 14 days) on the tenderization rate and protein changes in ten Martina Franca donkey striploins using a proteomic approach. During aging, a progressive decrease in share force, hardness, gumminess, and chewiness together with an increase in myofibril fragmentation index were observed. Proteolysis monitored by immunoblotting revealed a progressive degradation of desmin and fast troponin-T over time and an increase of their degradation products up to 14 days aging. Proteomics revealed by means of two-dimensional electrophoresis 37 protein spots corresponding to 15 proteins to change significantly by increasing aging time. These proteins belong to three pathways, these being "muscle contraction, structure, and associated proteins" (9 proteins); ii) "energy metabolism" (5 proteins); and iii) "chaperone" (1 protein). This study is the first to highlight the possible role of interconnected pathways in driving the final quality of donkey meat and predicting its texture.

The effects of quinoa protein Pickering emulsion (QPE) on the gel properties, protein structure and intermolecular interactions of myofibrillar protein (MP) gels were studied. Compared with the MP gels without QPE, the MP gels with 5.0%-7.5% added QPE showed significant increasing trends in storage modulus (G'), whiteness, gel strength and water holding capacity (WHC). The content of disulfide bonds in the gel increased with the addition of QPE and the disulfide bond conformation changed from gauche-gauche-gauche to gauche-gauche-trans. Moreover, the increase of hydrogen bonds after QPE addition confirmed the transformation from α-helix to β-sheet, as β-sheet structure was stabilized by interchain hydrogen bonds. The added QPE also enhanced the hydrophobic interaction and electrostatic interaction of MP gels. To conclude, the addition of 5.0%-7.5% QPE improved the intermolecular interactions and the structure stability of MP gels, and enhanced the gelation and WHC of MP gels.

Muscles sense internally generated and externally applied forces, responding to these in a coordinated hierarchical manner at different time scales. The center of the basic unit of the muscle, the sarcomeric M-band, is perfectly placed to sense the different types of load to which the muscle is subjected. In particular, the kinase domain (TK) of titin located at the M-band is a known candidate for mechanical signaling. Here, we develop the quantitative mathematical model that describes the kinetics of TK-based mechanosensitive signaling, and predicts trophic changes in response to exercise and rehabilitation regimes. First, we build the kinetic model for TK conformational changes under force: opening, phosphorylation, signaling and autoinhibition. We find that TK opens as a metastable mechanosensitive switch, which naturally produces a much greater signal after high-load resistance exercise than an equally energetically costly endurance effort. Next, in order for the model to be stable, give coherent predictions, in particular the lag following the onset of an exercise regime, we have to account for the associated kinetics of phosphate (carried by ATP), and for the non-linear dependence of protein synthesis rates on muscle fibre size. We suggest that the latter effect may occur via the steric inhibition of ribosome diffusion through the sieve-like myofilament lattice. The full model yields a steady-state solution (homeostasis) for muscle cross-sectional area and tension, and a quantitatively plausible hypertrophic response to training as well as atrophy following an extended reduction in tension.

Rabbit cardiac myosin contains fewer cysteine residues than the skeletal myosin (7 and 8.8 moles/10

After the discovery of troponin by Ebashi almost sixty years ago the field of striated muscle regulation has made significant progress. In the 1970’s the nascent troponin field gained momentum, including contributions by James D. Potter who established the stoichiometry of contractile proteins in the myofibril (Arch Biochem Biophys. 1974 Jun; 162(2):436-41. https://doi.org/10.1016/0003-9861(7490202-1)). This opened the door to refinement of competing models that described possible thick filament configurations. This study suggested the presence of one myosin per cross bridge and provided accurate calculations of the molar ratios of each protein - myosin: actin: tropomyosin: troponin T: troponin I: troponin C.

This was the first study to perform correlation analysis to determine the relationships between tertiary structural and rheological properties of the myofibrillar proteins (MPs) under ultrasound treatment (UT, frequency 20 kHz, power 500 W, intensity 30 %) for 0, 2, 4, 6, 8, and 10 min. The objectives of this study were to 1) characterize the changes of the small and large deformation rheological properties of MPs based on the tertiary structural changes; 2) determine relationships of rheological property, tertiary structural changes, and UT time. The results showed that UT could enhance the structural rigidity and the resistance of the system to permanent deformation observed in rheological tests. Besides, UT could unfold MPs, expose more hydrophobic amino acid residues to increase the surface hydrophobicity (S

Broiler chicks in the immediate posthatch handling period are exposed to thermal stress, with potentially harmful consequences for muscle growth and structure (e.g., less protein and more fat deposition). We addressed the effects of broiler chicks’ exposure to various ambient temperatures during the first 13 D posthatch on their performance, as well as on muscle development and structure, up to day 35. Body weight and pectoralis muscle growth were lower throughout the entire period in the high-heat-exposed chicks (39°C, Hot) and to a lesser extent in the mild-heat-exposed chicks (35°C, Hot Mild) than in the Control chicks that were raised under a commercial protocol. In the cold-exposed chicks (29° C, Cold), BW and pectoralis muscle absolute growth were similar to the Control group throughout the entire period. The lower body and muscle growth in the Hot and Hot Mild groups were reflected in a lower number of myonuclei expressing proliferating cell nuclear cell in pectoralis major muscle cross sections sampled on day 8, in the distribution of myofibers as the experiment progressed, and in mean myofiber diameter on day 35, whereas in the Cold group, these numbers exceeded that of the Control group. However, TUNEL assay revealed similar cell survival in all groups. Hematoxylin–eosin and Oil red O staining revealed the highest fat deposition in the pectoralis muscle derived from the Hot group, whereas lower fat deposition was observed in the Control Cold group. These results were corroborated by immunostaining for CCAAT/enhancer binding protein β in the pectoralis muscle, the levels of which were significantly higher in the Hot and Hot Mild groups on day 35 than in the Control group. Similar results were observed with Sirius red staining for collagen content in the pectoralis muscle. Together, the results imply long-term effects of chronic heat stress vs. cold stress in the early posthatch period on the broiler's body and muscle growth in general and myodegeneration of the pectoralis muscle in particular.

OBJECTIVE: Motor Unit Number Estimation (MUNE) methods, such as the recently developed MScanFit MUNE (MScan), may be valuable in tracking motor unit loss in ALS. Muscle Velocity Recovery Cycles (MVRCs) provide information about muscle membrane properties and can reveal disease-related changes. This study was undertaken to test the applicability of MScan to the anterior tibial muscle (TA) and to test whether the MVRCs could improve understanding of ALS pathophysiology.METHODS: Twenty-six ALS patients and 25 healthy controls were evaluated by quantitative electromyography, nerve conduction study and the two novel methods: MScan and MVRC; all in the TA and peroneal nerve.RESULTS: The estimated number of motor units for ALS patients (Median: 45, interquartile range: 28.5-76.5) was significantly lower than for the controls (117, 96.0-121.0) (P = 2.19 × 10-7). Unit size was increased only when amplitudes were expressed as percentage of CMAP. Of MVRC measurements, only relative refractory period was significantly abnormal in patients.CONCLUSION: MScanFit MUNE gives a sensitive and quantitative measure of loss of TA motor units in ALS. Muscle fiber membrane properties are mostly unaffected, despite substantial denervation, presumably due to collateral reinnervation.SIGNIFICANCE: MScan is suitable for detecting motor unit loss in TA. MVRCs do not provide new insights in ALS.

This study investigates the purification and biochemical characteristics of the protease secreted by Lactobacillus curvatus R5, which was isolated from Harbin dry sausages. The optimized fermentation conditions were fermentation time 36 h, initial pH 6 and fermentation temperature 37 °C. An extracellular protease was purified using ammonium sulfate precipitation, ion-exchange layer and gel filtration. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis showed that molecular weight of the purified protease was 45.3 kDa. Protease produced by L. curvatus R5 reached a higher relative protease activity at pH 6, 40 °C, and the purified protease exhibited pH and thermal stability at pH 6 and 40 °C. The microbial protease activity can be inhibited by ethylene diamine tetraacetic acid disodium salt (EDTA). The Vmax and Km of the protease were 53 mg/min and 15.9 mg/mL, respectively. SDS-PAGE reflects the ability of the protease to hydrolyse myofibrillar protein and sarcoplasmic protein, especially on myosin heavy chain, actin, myosin light chain and phosphorylase. The 3D structure and the Ramachandran plot of L. curvatus R5 protease was obtained by homology modelling. The Ramachandran plot analysis revealed that the purified protease was composed of 366 amino acids, and its residues in favoured, allowed, generously allowed and disallowed regions were 84.6%, 11.3%, 3.2% and 0.9% residues, respectively. Molecular docking showed that the substrate actin bound to the protease active site by hydrogen bonding and hydrophobic interaction. This research provides a basis for understanding the enzymatic properties of L. curvatus R5 protease. In conclusion, L. curvatus R5 can be used as a starter culture or protease-producing strain to inoculate Harbin dry sausages.

Effects of two typical comminution methods (shearing and blending) on structural and biochemical properties of silver carp surimi were comparatively investigated. Surimi myofibrils in striated appearance were progressively disrupted within 15 min of blending. The myofibrils were completely disintegrated after shearing for 5 min. Surface hydrophobicity of surimi increased and then gradually decreased (p 0.05) under shearing, while it continuously increased (p 0.05) under blending. As shearing time was extended, α-helical structure decreased and β-sheet structure increased simultaneously; surface active sulfhydryl content (SH) increased and then decreased (p 0.05); and intensity of myosin heavy chain (MHC) was gradually reduced. However, secondary structure, MHC intensity and SH were slightly changed as blending time extended. Ca

Specific ultrastructural anatomy of masticatory muscles is commonly referred to a general pattern assigned to striated muscles. Junctional feet consisting of calcium channels of the sarcoplasmic reticulum (i.e. the ryanodine receptors, RyRs) physically connected to the calcium channels of the t-tubules build triads within striated muscles. Functional RyRs were demonstrated in the nuclear envelopes of pancreas and of a skeletal muscle derived cell line, but not in muscle in situ. It was hypothesized that ryanodine receptors (RyRs) could also exist in the nuclear envelope in the masseter muscle, thus aiming at studying this by transmission electron microscopy. There were identified paired and consistent subsarcolemmal clusters of mitochondria, appearing as outpockets of the muscle fibers, usually flanking an endomysial microvessel. It was observed on grazing longitudinal cuts that the I-band-limited mitochondria were not strictly located in a single intermyofibrillar space but continued transversally over the I-band to the next intermyofibrillar space. It appeared that the I-band-limited transverse mitochondria participate with the column-forming mitochondria in building a rather incomplete mitochondrial reticulum of the masseter muscle. Subsarcolemmal nuclei presented nuclear envelope-associated RyRs. Moreover, t-tubules were contacting the nuclear envelope and they were seemingly filled from the perinuclear space. This could suggest that nucleoplasmic calcium could contribute to balance the cytosolic concentration via pre-built anatomical routes: (i) indirectly, via the RyRs of the nuclear envelope and (ii) directly via the communication of t-tubules and sarcoplasmic reticulum through the perinuclear space.

The effect and mechanisms of calcium chloride and citric acid addition on the acid method of processing tilapia protein isolates and surimi gels were studied. The alkaline method shows better gel quality than the acid method with lower yield and high lipid content. The addition of calcium chloride increased the breaking force to 494.56 g and citric acid helped reducing the lipid content by 85.85%, resulting in a higher yield of 75.36%. Gels with calcium maintained low levels of expressible moisture (2.30%), indicating a well-organised gel matrix. The decrease in -SH content and increase in ionic bonds content suggested increased formation of disulphide bonds and ionic bonds. Oscillatory dynamic measurement indicated that more heat resistant bonds were formed. The morphology of myofibril proteins studied by atomic force microscopy (AFM) showed that the height of the proteins in the calcium-added gels decreased 79.4%, which suggested that the protein structure was “suppressed”.

Beef colour is essential to consumer acceptability with dark muscle colours being problematic. Dark meat has less light scattering but the mechanisms are unknown. We hypothesise that three mechanisms are responsible for decreased light scattering in dark meat, namely (i) larger lateral separation of myofilaments, (ii) decreased optical protein density in the I-band and (iii) decreased denaturation of sarcoplasmic proteins. Nineteen beef longissimus thoracis muscles, divided into 'light', 'medium' and 'dark' colour groups, were assessed for light scattering by reflectance confocal microscopy, sarcomere length, and myofilament lattice spacing by small-angle X-ray diffraction. Dark muscles had a longer lattice spacing, shorter sarcomeres and wider muscle fibre diameters compared to lighter colour groups, indicating that the transverse spacing of muscle fibres that occurs post-mortem, with pH decline, is central to light scattering development. Dark muscles also had more degradation of the Z-disc and higher sarcoplasmic protein activities, which could impact on the optical density and contribute to lower light scattering.

Mitochondria represent the main source of ATP in skeletal muscle and mitochondria activity increases after muscle fiber depolarization. The regulation of mitochondrial function during contraction in skeletal muscle, however, is poorly understood. Skeletal muscle has a particular distribution of mitochondria where three distinct populations can be recognized. The most studied populations are the ones positioned deep into the myofibers between the myofibrils (intermyofibrillar mitochondria), and that located immediately beneath sarcolemma (subsarcolemmal mitochondria); a less studied population locates covering the myonuclei, as a continuation of the subsarcolemmal population. All mitochondria populations undergo fusion and fission events and intermyofibrillar mitochondria are interconnected; mitochondrial communication is necessary to maintain not only the energetic homeostasis of the muscle but its contractile function, as well. The mechanism supporting communication between subsarcolemmal and intermyofibrillar mitochondria is unknown. The recently described MCU complex of proteins has provided a new insight into the role of calcium as a regulator of mitochondrial function. Whether the different mitochondria populations have different calcium handling capacity and whether mitochondria Ca2+ has a role in energy transmission along the mitochondria network are intriguing issues that emerge when studying the link between electrical stimulation of the muscle fiber and the mitochondria metabolic output.

Movement of the myocardium can modify organ-level cardiac function and its molecular (crossbridge) mechanisms. This motion, which is defined by myocardial strain and strain rate (muscle shortening, lengthening, and the speed of these movements), occurs throughout the cardiac cycle, including during isovolumic periods. This review highlights how the left ventricular myocardium moves throughout the cardiac cycle, how muscle mechanics experiments provide insight into the regulation of forces used to move blood in and out of the left ventricle, and its impact on (and regulation by) crossbridge and sarcomere kinetics. We specifically highlight how muscle mechanics experiments explain how myocardial relaxation is accelerated by lengthening (strain rate) during late systole and isovolumic relaxation, a lengthening which has been measured in human hearts. Advancing and refining both in vivo measurement and ex vivo protocols with physiologic strain and strain rates could reveal important insights into molecular (crossbridge) kinetics. These advances could provide an improvement in both diagnosis and precise treatment of cardiac dysfunction.

Phosphorylation of cardiac sarcomeric proteins plays a major role in the regulation of the physiological performance of the heart. Phosphorylation of thin filament proteins, such as troponin I and T, dramatically affects calcium sensitivity of the myofiber and systolic and diastolic functions. Phosphorylation of the regulatory protein tropomyosin (Tpm) results in altered biochemical properties of contraction; however, little is known about the physiological effect of Tpm phosphorylation on cardiac function. To address the in vivo significance of Tpm phosphorylation, here we generated transgenic mouse lines having a phosphomimetic substitution in the phosphorylation site of α-Tpm (S283D). High expression of Tpm S283D variant in one transgenic mouse line resulted in an increased heart:body weight ratio, coupled with a severe dilated cardiomyopathic phenotype resulting in death within 1 month of birth. Moderate Tpm S283D mice expression in other lines caused mild myocyte hypertrophy and fibrosis, did not affect lifespan, and was coupled with decreased expression of extracellular signal-regulated kinase 1/2 kinase signaling. Physiological analysis revealed that the transgenic mice exhibit impaired diastolic function, without changes in systolic performance. Surprisingly, we observed no alterations in calcium sensitivity of the myofibers, cooperativity, or calcium-ATPase activity in the myofibers. Our experiments also disclosed that casein kinase 2 plays an integral role in Tpm phosphorylation. In summary, increased expression of pseudo-phosphorylated Tpm impairs diastolic function in the intact heart, without altering calcium sensitivity or cooperativity of myofibers. Our findings provide the first extensive in vivo assessment of Tpm phosphorylation in the heart and its functional role in cardiac performance.

Background Age-related decline in skeletal muscle mass is at least partly attributed to anabolic resistance to food intake. Resistance exercise sensitizes skeletal muscle tissue to the anabolic properties of amino acids. Objective The present study assessed protein digestion and amino acid absorption kinetics, whole-body protein balance, and the myofibrillar protein synthetic response to ingestion of different amounts of protein during recovery from resistance exercise in older men. Methods Forty-eight healthy older men [mean ± SEM age: 66 ± 1 y; body mass index (kg/m2): 25.4 ± 0.3] were randomly assigned to ingest 0, 15, 30, or 45 g milk protein concentrate after a single bout of resistance exercise consisting of 4 sets of 10 repetitions of leg press and leg extension and 2 sets of 10 repetitions of lateral pulldown and chest press performed at 75–80% 1-repetition maximum. Postprandial protein digestion and amino acid absorption kinetics, whole-body protein metabolism, and myofibrillar protein synthesis rates were assessed using primed, continuous infusions of l-[ring-2H5]-phenylalanine, l-[ring-2H2]-tyrosine, and l-[1-13C]-leucine combined with ingestion of intrinsically l-[1-13C]-phenylalanine and l-[1-13C]-leucine labeled protein. Results Whole-body net protein balance showed a dose-dependent increase after ingestion of 0, 15, 30, or 45 g of protein (0.015 ± 0.002, 0.108 ± 0.004, 0.162 ± 0.008, and 0.215 ± 0.009 μmol Phe · kg−1 · min−1, respectively; P

The effects of thawing methods (refrigeration thawing (RT, 4 °C), water immersion thawing (WT, 18 °C), vacuum thawing (VT, 25 °C), ultrasonic thawing (UT, 20 °C) and microwave thawing (MT)) on the conformation and gel qualities of myofibrillar protein (MP) obtained from porcine longissimus muscle were investigated. The results showed that MP conformation and gel qualities of porcine longissimus muscles by VT and UT were insignificantly changed compared to fresh meat (FM). A significant decrease in free amino groups of MP from MT illustrated that MT induced protein aggregation and oxidation (P

Rationale Mutations in the gene encoding the sarcomeric protein cardiac myosin-binding protein C (cMyBP-C) are a leading cause of hypertrophic cardiomyopathy (HCM). Mouse models targeting cMyBP-C and use of recombinant proteins have been effective in studying its roles in contractile function and disease. Surprisingly, while the N-terminus of cMyBP-C is important to regulate myofilament binding and contains many HCM mutations, an incorrect sequence, lacking the N-terminal 8 amino acids has been used in many studies. Objectives To determine the N-terminal cMyBP-C sequences in ventricles and investigate the roles of species-specific differences in cMyBP-C on myofilament binding. Methods and results We determined cMyBP-C sequences in mouse and human by inspecting available sequence databases. N-terminal differences were confirmed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Cosedimentation assays with actin or myosin were used to examine binding in mouse, human and chimeric fusion proteins of cMyBP-C. Time-resolved FRET (TR-FRET) with site-directed probes on cMyBP-C was employed to measure structural dynamics. LC-MS/MS supported the sequencing data that mouse cMyBP-C contains an eight-residue N-terminal extension (NTE) not found in human. Cosedimentation assays revealed that cardiac myosin binding was strongly influenced by the presence of the NTE, which reduced binding by 60%. 75% more human C0-C2 than mouse bound to myosin. Actin binding of mouse C0-C2 was not affected by the NTE. 50% more human C0-C2 than mouse bound to actin. TR-FRET indicates that the NTE did not significantly affect structural dynamics across domains C0 and C1. Conclusions Our functional results are consistent with the idea that cardiac myosin binding of N-terminal cMyBP-C is reduced in the mouse protein due to the presence of the NTE, which is proposed to interfere with myosin regulatory light chain (RLC) binding. The NTE is a critical component of mouse cMyBP-C, and should be considered in extrapolation of studies to cMyBP-C and HCM mechanisms in human.

To investigate the underlying mechanism of softening of ice-stored grass carp fillet, changes in assembly structure of myofibrillar proteins and potential candidates for regulating this change including myosin regulatory chain phosphorylation, heat shock proteins (Hsp27, Hsp90, αB-crystallin and UNC45) and endogenous protease activity were studied. Comparison of SDS-PAGE pattern of myofibrillar proteins treated with EDC crosslinking showed that thin filament experienced rapid disassembly within initial 8 h, followed by depolymerization of thick filament consisting of myosin, which further exacerbated the myofibril disorganization of fillets. Pearson coefficient analysis showed that UNC45, Hsp90, Hsp27 and αB-crystallin concentration and cathepsin B, D, L activities were significantly correlated with dissociated MHC and actin. Therefore, the significant correlation between shear force and dissociated MHC and actin clearly demonstrated that post mortem disassembly of myofibril filaments, which was regulated by endogenous proteolytic activity and loss of Hsp, contributed to the softening of ice-stored grass carp fillets.

This study aimed to determine the effect of frozen-then-chilled storage on free Ca2+, proteolytic enzyme activity of calpains and the proteasome, water-holding capacity and shear force of porcine longissimus thoracis et lumborum muscle. Pork loins were subjected to either chilled storage at 2 +/- 1 degrees C for 1, 2, 4, 6 and 9 days, or frozen then chilled storage (20 +/- 1 degrees C for 1 week followed by thawing overnight). Free Ca2+ increased with chilled storage in the non-frozen group. Frozen-then-chilled storage increased free Ca2+ concentration, followed by a faster decrease of calpain-1 activity and activation of around 50% of calpain-2. Proteasome activity was reduced by around 40% following freezing-thawing. Purge loss increased and water-holding capacity of myofibrils decreased in the frozen-thawed group, suggesting considerable denaturation of myofibrillar proteins. Shear force was not affected by freezing-thawing, and we speculate that the tenderizing effect of calpain activation was counteracted by loss of proteasome activity and substantial exudate loss.

This study was conducted to determine whether the application of high pressure processing (HPP) could prevent the process of rigor in fresh pork and to identify the physical and chemical changes occurring in the meat that are responsible for this effect. Pressure (0–400 MPa) and time (10 min) were applied to the pork loin and enzyme samples. The effects of treatment on shear force, myofibrillar fragmentation, enzyme activity and meat texture were measured. The results showed that, compared with untreated samples, HPP treatment inhibited the occurrence of rigor in pork. The myofibrillar fragmentation indices of HPP treated samples were higher than those of control samples and increased linearly with increasing pressure. Application of HPP prevented the process of rigor at 0–300 MPa, reduced calpain activity and markedly decreased calpastatin activity, resulting in an increase of total calpains. Above 300 MPa, the mechanical action of pressure led to continuous prevention of rigor.

Made available in DSpace on 2018-12-11T17:20:42Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-11-01 The objective of this research was to study the effects of salt reduction and the application of ultrasound (nominal current of 600 W cm−2 for 10 min) on the physicochemical properties, the microstructure and the sensory acceptance of restructured cooked ham. Four treatments with reduced salt including one with the application of ultrasound (1.5, 1.12, 0.75 and 0.75% salt + ultrasound) were produced. The treatment with 0.75% salt provided a reduction of about 30% in the sodium content. The use of ultrasound decreased the Total Fluid Release and increased the hardness. For lightness, the sample with 0.75% salt with the application of ultrasound did not differ from the control at day zero of storage. The use of ultrasound increased redness too. The ultrasound treatment caused micro fissures on the myofibrils. The sensory acceptance of restructured cooked ham with 0.75% of salt was improved with ultrasound applied. The ultrasound showed good potential for use in the production of healthier meat products. São Paulo State University – UNESP Institute of Biosciences Humanities and Exact Sciences – IBILCE Food Engineering and Technology Department - Meat and Meat Products Laboratory, Campus São José do Rio Preto, Cristovão Colombo St. 2265 Federal Institute of São Paulo – IFSP, Campus Barretos, C-1 Avenue 250 Faculty of Food Engineering University of Campinas Department of Food Technology, Monteiro Lobato St 80 São Paulo State University – UNESP Institute of Biosciences Humanities and Exact Sciences – IBILCE Food Engineering and Technology Department - Meat and Meat Products Laboratory, Campus São José do Rio Preto, Cristovão Colombo St. 2265

Mussel myofibrillar proteins (MMP) suspensions (10.6% ± 0.5%, w/v) were treated by high-pressure homogenization (HPH) at 0 (control), 20, 40, 60, 80 or 100 MPa for 3 cycles. Particle size distribution, zeta potential, solubility, water and oil holding capacity, emulsifying, foaming properties, secondary structure, free sulfhydryl and surface hydrophobicity of the obtained suspensions were analyzed. The results showed that functional properties of MMP significantly (P 0.05) improved after HPH treatment. Absolute zeta potential, emulsifying activity index, emulsion stability index, foaming ability and foaming stability increased by 23.64 mV, 14.99 m

Oxidants are produced endogenously and can react with and thereby post-translationally modify target proteins. They have been implicated in the redox regulation of signal transduction pathways conferring protection, but also in mediating oxidative stress and causing damage. The difference is that in scenarios of injury the amount of oxidants generated is higher and/or the duration of oxidant exposure sustained. In the cardiovascular system, oxidants are important for blood pressure homeostasis, for unperturbed cardiac function and also contribute to the observed protection during ischemic preconditioning. In contrast, oxidative stress accompanies all major cardiovascular pathologies and has been attributed to mediate contractile dysfunction in part by inducing oxidative modifications in myofilament proteins. However, the proportion to which oxidative modifications of contractile proteins are beneficial or causatively mediate disease progression needs to be carefully reconsidered. These antithetical aspects will be discussed in this review with special focus on direct oxidative post-translational modifications of myofilament proteins that have been described to occur in vivo and to regulate actin-myosin interactions in the cardiac myocyte sarcomere, the methodologies for detection of oxidative post-translational modifications in target proteins and the feasibility of antioxidant therapy strategies as a potential treatment for cardiac disorders.