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SummaryCellulose is an abundant cell wall component of land plants. It is synthesized from UDP-activated glucose molecules by cellulose synthase, a membrane-integrated processive glycosyltransferase. Cellulose synthase couples the elongation of the cellulose polymer with its translocation across the plasma membrane. Here, we present substrate and product-bound cryogenic electron microscopy structures of the homotrimeric cellulose synthase isoform-8 (CesA8) from hybrid aspen (poplar). UDP-glucose binds to a conserved catalytic pocket adjacent to the entrance to a transmembrane channel. The substrate’s glucosyl unit is coordinated by conserved residues of the glycosyltransferase domain and amphipathic interface helices. Site-directed mutagenesis of a conserved gating loop capping the active site reveals its critical function for catalytic activity. Molecular dynamics simulations reveal prolonged interactions of the gating loop with the substrate molecule, particularly across its central conserved region. These transient interactions likely facilitate the proper positioning of the substrate molecule for glycosyl transfer and cellulose translocation.HighlightsCryo-EM structures of substrate and product bound poplar cellulose synthase provide insights into substrate selectivitySite directed mutagenesis signifies a critical function of the gating loop for catalysisMolecular dynamics simulations support persistent gating loop – substrate interactionsGating loop helps in positioning the substrate molecule to facilitate cellulose elongationConserved cellulose synthesis substrate binding mechanism across the kingdoms

Higher plants synthesize cellulose using membrane-bound, six-lobed cellulose synthase complexes, each lobe containing trimeric cellulose synthases (CESAs). Although molecular biology reports support heteromeric trimers composed of different isoforms, a homomeric trimer was reported for in vitro studies of the catalytic domain of CESA1 of

Effective diagnosis of disease and its progression can be aided by 19 F magnetic resonance imaging (MRI) techniques. Specifically, the inherent sensitivity of the spin-lattice relaxation time (T1 ) of 19 F nuclei to oxygen partial pressure makes 19 F MRI an attractive non-invasive approach to quantify tissue oxygenation in a spatiotemporal manner. However, there are only few materials with the adequate sensitivity to be used as oxygen-sensitive 19 F MRI agents at clinically relevant field strengths. Motivated by the limitations in current technologies, we report highly fluorinated monomers that provide a platform approach to realize water-soluble, partially fluorinated copolymers as 19 F MRI agents with the required sensitivity to quantify solution oxygenation at clinically relevant magnetic field strengths. The synthesis of a systematic library of partially fluorinated copolymers enabled a comprehensive evaluation of copolymer structure-property relationships relevant to 19 F MRI. The highest-performing material composition demonstrated a signal-to-noise ratio that corresponded to an apparent 19 F density of 220 mm, which surpasses the threshold of 126 mm 19 F required for visualization on a three Tesla clinical MRI. Furthermore, the T1 of these high performing materials demonstrated a linear relationship with solution oxygenation, with oxygen sensitivity reaching 240×10-5 mmHg-1 s-1 . The relationships between material composition and 19 F MRI performance identified herein suggest general structure-property criteria for the further improvement of modular, water-soluble 19 F MRI agents for quantifying oxygenation in environments relevant to medical imaging.

Cellulose synthase (CESA) polymerizes glucose into β-1,4-glucan chains that assemble to form cellulose microfibrils. Cellulose is the most abundant natural polymer in the world and a major structural component of the plant cell wall. An understanding of cellulose synthesis in plants is crucial for advancement in biofuels research and requires high-resolution 3D CESA structures. However, the determination of 3D structures of plant CESAs and their specific hierarchical arrangement into cellulose synthesis complexes (CSCs) has been a challenge. The prediction of CESA structures using computational methods presents a challenge due to poor sequence homology with resolved structures, long sequence, and structural complexity due to a mixture of globular, transmembrane, and intrinsically disordered regions. Herein, we present a 3D atomic-resolution model of a full-length (974-aa) cotton CESA (GhCESA1) structure using a variety of computational techniques with a reasonable ProSA-web z-score of − 8.32 (PDB available in SI). The overall fold of the CESA model indicates that there are similarities to BcsA bacterial cellulose synthase, such as the transmembrane topology and the internalization of conserved catalytic motifs. The plant-specific regions (CSR, P-CR, and N-term) fold into distinct subdomains, indicating the importance of these regions in CESA assembly into plant CSCs. We further examined possible assemblies of CESA monomers forming trimers and 18-mer CSCs, and we compared our results to those obtained by freeze fracture transmission electron microscopy. We observed that there are numerous competing ways in which CESAs may be arranged into homotrimers and CSCs. Our predicted structure can be used to probe CESA structure–activity relationships, select and subsequently test possible mutants, and investigate CESA aggregation into CSCs and microfibril formation to optimize biomass properties.

Sub-Saharan Africa (SSA) is experiencing an increasing prevalence of young children being overweight and obese. Many feeding and physical activity-related behaviours are established at home during preschool years, yet the precise factors that contribute to preschool overweight and obesity have not been fully elucidated. This review aims to identify factors in the home environment associated with overweight and or obesity in preschool children in SSA. Ovid MEDLINE, EMBASE, CINAHL, Scopus, Web of Science, Africa Journals Online (AJOL) and the African Index Medicus databases were systematically searched for qualitative and quantitative studies published between 2000 and 2021. Eleven studies (ten quantitative, one qualitative) met the inclusion criteria. Overall, the results highlight the paucity of studies exploring factors in the home environment associated with overweight and obesity in preschool children in Sub-Saharan Africa. The home food environment and maternal BMI appear to be important factors associated with overweight and obesity in preschool children; however, the information for all other factors explored remains unclear due to the lack of evidence. For successful obesity prevention and treatment interventions to be developed, more research in this area is required to understand how different aspects of the home environment contribute to overweight and obesity in preschool Sub-Saharan African children.

Significance The investigation of protein-based materials has provided a better understanding of living systems and has led to the development of ubiquitous modern technologies. Within this context, unique cephalopod proteins called reflectins have exhibited promise for biophotonics and bioelectronics applications, but the exploration of reflectins as materials has been hindered by an incomplete understanding of their structures and properties. Here, we resolve the molecular-level structure of a model reflectin variant, establish a straightforward approach to controlling the assembly of this protein, and describe a correlation between its structural characteristics and light-manipulating properties. Taken together, our findings advance current understanding of reflectin-based materials, provide insight into the color-changing capabilities of cephalopods, and afford new opportunities in biochemistry, cellular biology, bioengineering, and optics., Naturally occurring and recombinant protein-based materials are frequently employed for the study of fundamental biological processes and are often leveraged for applications in areas as diverse as electronics, optics, bioengineering, medicine, and even fashion. Within this context, unique structural proteins known as reflectins have recently attracted substantial attention due to their key roles in the fascinating color-changing capabilities of cephalopods and their technological potential as biophotonic and bioelectronic materials. However, progress toward understanding reflectins has been hindered by their atypical aromatic and charged residue-enriched sequences, extreme sensitivities to subtle changes in environmental conditions, and well-known propensities for aggregation. Herein, we elucidate the structure of a reflectin variant at the molecular level, demonstrate a straightforward mechanical agitation-based methodology for controlling this variant’s hierarchical assembly, and establish a direct correlation between the protein’s structural characteristics and intrinsic optical properties. Altogether, our findings address multiple challenges associated with the development of reflectins as materials, furnish molecular-level insight into the mechanistic underpinnings of cephalopod skin cells’ color-changing functionalities, and may inform new research directions across biochemistry, cellular biology, bioengineering, and optics.

Effective diagnosis of disease and its progression can be aided by

We report herein on the use of all-atom molecular dynamics simulations to investigate the solvation environment of C60 and four C60-derived fullerenes immersed in a variety of aromatic solvents. Utilizing a recently developed solvation shell analysis technique that quantifies the spatial relationships between fullerenes and solvent on a molecular level, we show that the number of fullerene substituents and solvent chemistry are crucial determinants of the solvation shell structure and thus fullerene solvation behavior. Specifically, it is shown for the derivatives investigated that the number of fullerene substituents is more critical to solvation behavior than the substituent chemistry.

Nanoparticles (NPs) play increasingly important roles in nanotechnology and nanomedicine in which nanoparticle surface chemistry allows for control over interactions with other nanoparticles and biomolecules. In particular, for applications in drug and gene delivery, a fundamental understanding of the NP-nucleic acid interface allows for development of more efficient and effective nanoparticle carriers. Computational modeling can provide insights of processes occurring at the inorganic NP-nucleic interface in detail that is difficult to access by experimental methods. With recent advances such as the use of graphics processing units (GPUs) for simulations, computational modeling has the potential to give unprecedented insight into inorganic-biological interfaces via the examination of increasingly large and complex systems. In this Topical Review, we briefly review computational methods relevant to the interactions of inorganic NPs and nucleic acids and highlight recent insights obtained from various computational methods that were applied to studies of inorganic nanoparticle-nanoparticle and nanoparticle-nucleic acid interfaces.

Collagen type I crosslink type and prevalence can be influenced by age, tissue type, and health; however, the role that crosslink chemical structure plays in mechanical behavior is not clear. Molecular dynamics simulations of ~65-nm-long microfibril units were used to predict how difunctional (deH-HLNL and HLKNL) and trifunctional (HHL and PYD) crosslinks respond to mechanical deformation. Low- and high-strain stress-strain regions were observed, corresponding to crosslink alignment. The high-strain elastic moduli were 37.7, 37.9, 39.9, and 42.4 GPa for the HLKNL, deH-HLNL, HHL, and PYD-crosslinked models, respectively. Bond dissociation analysis suggests that PYD is more brittle than HHL, with deH-HLNL and HLKNL being similarly ductile. These results agree with the tissues in which these crosslinks are found (e.g., deH-HLNL/HLKNL in developing tissues, HHL in mature skin, and PYD in mature bone). Chemical structure-function relationships identified for these crosslinks can aid the development of larger-scale models of collagenous tissues and materials.

Ligand-functionalized inorganic nanoparticles, also known as monolayer-protected nanoparticles, offer great potential as vehicles for in vivo delivery of drugs, genes, and other therapeutics. These nanoparticles offer highly customizable chemistries independent of the size, shape, and functionality imparted by the inorganic core. Their success as drug delivery agents depends on their interaction with three major classes of biomolecules: nucleic acids, proteins, and membranes. Here, the authors discuss recent advances and open questions in the field of nanoparticle ligand design for nanomedicine, with a focus on atomic-scale interactions with biomolecules. While the importance of charge and hydrophobicity of ligands for biocompatibility and cell internalization has been demonstrated, ligand length, flexibility, branchedness, and other properties also influence the properties of nanoparticles. However, a comprehensive understanding of ligand design principles lies in the cost associated with synthesizing and characterizing diverse ligand chemistries and the ability to carefully assess the structural integrity of biomolecules upon interactions with nanoparticles.

Subfailure matrix injuries such as sprains and strains account for a considerable portion of ligament and tendon pathologies. In addition to the lack of a robust biological healing response, these types of injuries are often characterized by seriously diminished matrix biomechanics. Recent work has shown nanosized particles, such as nanocarbons and nanocellulose, to be effective in modulating cell and biological matrix responses for biomedical applications. In this article, we investigate the feasibility and effect of using high stiffness nanostructures of varying size and shape as nanofillers to mechanically reinforce damaged soft tissue matrices. To this end, nanoparticles (NPs) were characterized using atomic force microscopy and dynamic light scattering techniques. Next, we used a uniaxial tensile injury model to test connective tissue (porcine skin and tendon) biomechanical response to NP injections. After injection into damaged skin and tendon specimens, the NPs, more notably nanocarbons in skin, led to an increase in elastic moduli and yield strength. Furthermore, rat primary patella tendon fibroblast cell activity evaluated using the metabolic water soluble tetrazolium salt assay showed no cytotoxicity of the NPs studied, instead after 21 days nanocellulose-treated tenocytes exhibited significantly higher cell activity when compared with nontreated control tenocytes. Dispersion of nanocarbons injected by solution into tendon tissue was investigated through histologic studies, revealing effective dispersion and infiltration in the treated region. Such results suggest that these high modulus NPs could be used as a tool for damaged connective tissue repair.

The anterior cruciate ligament (ACL) is important for knee stability and kinematics. It is also the most com- monly injured ligament of the knee and due to its poor healing potential, severe damage warrants surgical intervention in- cluding complete replacement. Therefore, investigators have begun to pursue new techniques and devices for the repair, regeneration, and replacement of the ACL. These options involve the use of mechanically functional grafts that are designed to increase implant stability in order to withstand normal mechanical loads (while promoting ligament develop- ment in some cases). This article presents background on the ACL and its replacement, novel replacement approaches utilizing a variety of materials, and recent patent coverage.

Collagen type I is a structural protein that provides tensile strength to tendons and ligaments. Type I collagen molecules form collagen fibers, which are viscoelastic and can therefore store energy elastically via molecular elongation and dissipate viscous energy through molecular rearrangement and fibrillar slippage. The ability to store elastic energy is important for the resiliency of tendons and ligaments, which must be able to deform and revert to their initial lengths with changes in load. In an earlier paper by one of the present authors, molecular modeling was used to investigate the role of mineralization upon elastic energy storage in collagen type I. Their collagen model showed a similar trend to their experimental data but with an over-estimation of elastic energy storage. Their simulations were conducted in vacuum and employed a distance-dependent dielectric function. In this study, we performed a re-evaluation of Freeman and Silver's model data incorporating the effects of explicit solvation and water infiltration, in order to determine whether the model data could be improved with a more accurate representation of the solvent and osmotic effects. We observed an average decrease in the model's elastic energy storage of 45.1%±6.9% in closer proximity to Freeman and Silver's experimental data. This suggests that although the distance-dependent dielectric implicit solvation approach was favored for its increased speed and decreased computational requirements, an explicit representation of water may be necessary to more accurately model solvent interactions in this particular system. In this paper, we discuss the collagen model described by Freeman and Silver, the present model building approach, the application of the present model to that of Freeman and Silver, and additional assumptions and limitations.

The anterior cruciate ligament (ACL) is important for knee stability and kinematics. Unfortunately, it is also the most commonly injured ligament of the knee and due to its poor healing potential, severe damage warrants surgical intervention including complete replacement. Current ACL replacements have a host of limitations that prevent their extensive use. Thus, investigators have begun to pursue tissue engineering techniques to create new options for the repair, regeneration, and replacement of the ACL. These options involve devices that are mechanically functional tissue engineered scaffolds and as such are designed to withstand normal mechanical loads while promoting ligament development. This article presents background on the ACL and its replacement, novel replacement approaches utilizing natural polymers, synthetic polymers, and natural tissues, and recent patent coverage.

The impact of extraction methodology and polarity of extraction solvents on the assay of phenolic compounds was investigated using parsley (Petroselinum crispum) flakes as a model substrate. This systematic study was undertaken to address substantial variations in the extraction procedures, solvents and conditions as described in the recent literature. Five different extraction procedures [shaking, vortex mixing, sonication, stirring and pressurized liquid extraction (PLE)] and three different solvents (methanol, ethanol and acetone), with five different solvent to water ratios per solvent, were used for extraction. Extracts were analyzed for phenolic content by high-performance liquid chromatography and Folin–Ciocalteu assays. The yields of phenolic compounds extracted with a pressurized liquid extractor were comparable to or better than those of four classical extraction procedures. Optimum extraction efficiency with PLE was obtained when extractions were performed with four extraction cycles using ethanol–water (50:50, v/v). The amount of apiin (4,5,7-trihydroxyflavone 7-apiosylglucoside) and malonylapiin (apigenin malonylapiosylglucoside) isolated from parsley varied with the composition of extraction solvent. Apiin extractability was found to be a maximum when the solvent (ethanol, methanol or acetone) to water ratio was 30:70 (v/v), whereas higher amounts of malonylapiin were isolated with a reverse solvent to water ratio (70:30, v/v). Malonylapiin was not detected when parsley samples were extracted with organic solvent to water ratios of 10:90 (v/v) and 30:70 (v/v). Published in 2006 by John Wiley & Sons, Ltd

Determination of niacin in fortified infant and dairy products has been accomplished using a variety of analytical liquid chromatography (LC) methods. Applications of these LC techniques to other food matrices suffer due to the presence of endogenous absorbing peaks at 260 nm that co-chromatograph with the nicotinic acid and nicotinamide vitamers. We have successfully adapted the LC method of Woollard and Indyk for the determination of nicotinamide in reference and commercial cereal products. Unbound nicotinamide in fortified cereal was extracted with 0.6M tri-chloroacetic acid and chromatographed on a C18 reversed-phase column using a mobile phase of 75% methanol and water (pH 2.8, with formic acid) with sodium dioctylsulphosuccinate (0.1%) as the ion-pairing agent. Using Spectral Analysis ChromQuest software, a three-dimensional view showed only nicotinamide under the LC peak. Similarity index spectral matches of nicotinamide standard and the LC peak were ≈100%, indicating the absence of interf...

Collagen type I is an extracellular matrix protein found in connective tissues such as tendon, ligament, bone, skin, and the cornea of the eyes, where it functions to provide tensile strength; it also serves as a scaffold for cells and other extracellular matrix components. A single collagen type I molecule is composed of three amino acid chains that form a triple helix for most of the molecule's length; non-triple-helical extensions called N- and C-telopeptides are located at the amino/N-terminal and carboxy/C-terminal ends of the molecule, respectively. In two of the three chains, the C-telopeptide has been reported to possess a hair-pin/hook conformation, while the three N-telopeptides display a more extended structure. These telopeptides are crucial for the formation of enzymatic covalent crosslinks that form in collagens near their N- and C-ends. Such crosslinks provide structural integrity, strength, and stiffness to collagenous tissues. However, deformation mechanisms of N- and C-crosslinks and functional roles for the N- and C-telopeptide conformations are not yet well known. By performing molecular dynamics simulations, we demonstrated that two dehydro-hydroxylysino-norleucine crosslinks, positioned at the N- and C-crosslinking sites, exhibited a two-stage response to the mechanical deformation of their parent molecules. We observed that the N-crosslink served as the first responder to mechanical deformation, followed by the C-crosslink. The results of our simulations suggest a mechanical recruitment mechanism for N- and C-crosslinks. Understanding this mechanism will be crucial for the development of larger-scale predictive models of the mechanical behavior of native collagenous tissues, engineered tissues, and collagen-based materials.

Collagen type I fiber-based scaffolds for anterior cruciate ligament (ACL) replacement were evaluated for their mechanical properties and their ability to promote cellular proliferation. Prior to scaffold formation, two crosslinking methods were investigated on individual reconstituted collagen type I fibers, ultraviolet radiation, and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). Crosslinking with EDC for 4 hr yielded mechanical properties similar to the human ACL; therefore, scaffold crosslinking was done with EDC for 4 hr. A braid-twist scaffold design was used, and scaffolds were left uncrosslinked, crosslinked after the addition of gelatin, or crosslinked without gelatin. The ultimate tensile strength, Young's modulus, and viscoelastic properties of the scaffolds were then evaluated. In order to assess cellular response on the scaffolds, primary rat ligament fibroblast cells were seeded upon the scaffolds. Cell activity was evaluated at days 7, 14, and 21 using a Cell Titer 96(®) AQueous One Solution Cell Proliferation Assay (MTS Assay). The mechanical testing results showed that among the three scaffold groups, the crosslinked scaffolds without gelatin displayed an ultimate tensile strength, Young's modulus, and viscoelastic properties that were closest to the human ACL. Improvements are still desired to enhance the mechanical compliance and ductility of these scaffolds. Cell activity was observed on all cell-seeded scaffolds by day 7, but by day 21 only the crosslinked scaffolds without gelatin displayed increased cellular activity compared with the negative controls. Although improvement is still needed, the results suggest that these scaffolds have the potential to contribute toward an ACL replacement strategy.

A new mathematical model is presented to describe the elastic and viscoelastic properties of a single collagen fiber. The model is formulated by accounting for the mechanical contribution of the collagen fiber's main constituents: the microfibrils, the interfibrillar matrix and crosslinks. The collagen fiber is modeled as a linear elastic spring, which represents the mechanical contribution of the microfibrils, and an arrangement in parallel of elastic springs and viscous dashpots, which represent the mechanical contributions of the crosslinks and interfibrillar matrix, respectively. The linear elastic spring and the arrangement in parallel of elastic springs and viscous dashpots are then connected in series. The crosslinks are assumed to gradually break under strain and, consequently, the interfibrillar is assumed to change its viscous properties. Incremental stress relaxation tests are conducted on dry collagen fibers reconstituted from rat tail tendons to determine their elastic and viscoelastic properties. The elastic and total stress-strain curves and the stress relaxation at different levels of strain collected by performing these tests are then used to estimate the parameters of the model and evaluate its predictive capabilities.

Among the five ligaments in the knee the anterior cruciate ligament (ACL) is among the most important for stability and also the most commonly injured. Due to a lack of vascularization, the ACL has poor healing potential, therefore moderate to severe damage warrants medical intervention. Ligaments are complex, highly organized tissues; they are longitudinally arranged with a great deal of order that begins at the molecular level and carries through to the tissue level. The components of the ligament and their location and orientation heavily influence the tissue's mechanical behavior. ACL replacements have faced a variety of limitations that prevented their extensive use, including implant fatigue or fraying of the device. In the face of these problems, investigators have begun to examine a variety of matrix based techniques to create options for ACL repair, replacement, and regeneration. This article will discuss ACL structure and mechanics, past replacement options and their limitations, and recent tissue engineered options for ACL repair. These techniques employ a wide variety of designs, materials, and methods to heal damaged ACL tissue or regenerate lost tissue in order to regain full ACL strength and mechanics.