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Graphical abstract Highlights • Peptide motif sequences of the first helix of osteocalcin (OC) is synthesized in acidic and amidic forms. • An effective dosage of peptides is advanced alkaline phosphatase activity. • The amidated peptide stimulated RNA expression of collagen type I and osteopontin. • Mineral deposition is also considerably increased in the presence of amidated peptide. • The osteocalcin mimetic peptides can be utilized as a bioactive inducer for hard tissue regeneration. Abstract A natural peptide motif in the first helix of osteocalcin (OCN) is used to promote nucleation and crystallization of hydroxyapatite (HA) in hard tissue. The capability of osteocalcin mimetic peptides to induce osteogenic activity of osteoblast cells leading to in-vitro mineralization is demonstrated. An osteocalcin-derived peptide consisting of thirteen amino acids is synthesized in both acidic (OSC) and amidic (OSN) forms and added into the human osteoblast-like cells (MG63) culture. The viability, proliferation, alkaline phosphatase activity, HA deposition and osteogenic gene expression by osteoblast cells are evaluated. It is revealed that the addition of 100 μg/ml of peptides enhances the proliferation rate and total protein content of osteoblast cells. Alkaline phosphatase activity is significantly higher in the presence of peptides which in turn stimulated RNA expression of collagen type I and osteopontin in a phosphate-dependent manner. Alizarin red staining and calcium content measurement show that mineral deposition is considerably increased. Ultrastructural characterization of MG63 cultures confirms the crystalline nature and chemical composition of HA mineral formation in the presence of peptides. It is confirmed that the osteocalcin-derived peptide, particularly in amidic form (OSN), is able to act as a bioactive inducer of mineralization process and hence accelerating bone tissue regeneration. [ABSTRACT FROM AUTHOR]

Genesis of natural biocomposite-based materials, such as bone, cartilage, and teeth, involves interactions between organic and inorganic systems. Natural biopolymers, such as peptide motif sequences, can be used as a template to direct the nucleation and crystallization of hydroxyapatite (HA). In this study, a natural motif sequence consisting of 13 amino acids present in the first helix of osteocalcin was selected based on its calcium binding ability and used as substrate for nucleation of HA crystals. The acidic (acidic osteocalcin-derived peptide (OSC)) and amidic (amidic osteocalcin-derived peptide (OSN)) forms of this sequence were synthesized to investigate the effects of different C termini on the process of biomineralization. Electron microscopy analyses show the formation of plate-like HA crystals with random size and shape in the presence of OSN. In contrast, spherical amorphous calcium phosphate is formed in the presence of OSC. Circular dichroism experiments indicate conformational changes of amidic peptide to an open and regular structure as a consequence of interaction with calcium and phosphate. There is no conformational change detectable in OSC. It is concluded that HA crystal formation, which only occurred in OSN, is attributable to C-terminal amidation of a natural peptide derived from osteocalcin. It is also proposed that natural peptides with the ability to promote biomineralization have the potential to be utilized in hard tissue regeneration. [ABSTRACT FROM AUTHOR]

Abstract: The industrial utilization of savinase is mainly constrained by its stability limitations. In the present study, the irreversible thermoinactivation of savinase has been evaluated at 70°C, and various possible mechanisms for irreversible thermoinactivation of savinase were examined. The main process seemed to be autodigestion of savinase at higher temperatures. To improve the thermal stability of the enzyme, the effect of two co-solvents (sorbitol and trehalose) on the enzyme''s activity and stability was investigated. Both osmolytes prevented the autolysis of savinase at 70°C without inactivating the enzyme; furthermore, the structural and kinetic stabilities of the enzyme increased in the presence of additives. [Copyright &y& Elsevier]

Abstract: Bacillus sp. KR-8104 was selected from a set of 18 bacteria strains isolated from soil samples and screened for production of amylase. The maximum productivity obtained at pH 5–6 and 60–65h after cultivation in production medium. New extracellular Ca-independent α-amylase was highly purified using ion exchange and hydrophobic interaction chromatography, which showed a single band with an apparent molecular weight of 59kDa by SDS-PAGE. This enzyme is active in a wide pH range with its maximum activity at low pH values (4.0–6.0) and has the 90% of its maximum activity at pH 3.5. The α-amylase is optimally active at 75–80°C. The presence or absence of Ca2+ and EDTA did not affect enzyme activity and thermal stability. [Copyright &y& Elsevier]

• In vivo-evaluated 5-ASA-HbNPs as a colon-specific prodrug for treatment of experimental colitis. • HbNPs conjugation is an efficient way to increase the therapeutic effect and to reduce the adverse effects of 5-ASA. • HbNPs acted as not only a pro-moiety but also a therapeutically active agent. • Mucoadhesive property of this colonic nano-delivery system contributed to the treatment of IBD. The oral formulation design for colon-specific drug delivery brings some therapeutic benefits in the ulcerative colitis treatment. We recently reported the specific delivery of hemoglobin nanoparticles-conjugating 5-aminosalicylic acid (5-ASA-HbNPs) to the inflamed site. In the current study, the therapeutic effect of the 5-ASA-HbNPs formulation was confirmed in vivo. This evaluation of 5-ASA-HbNPs not only shows longer colonic retention time due to adhesive properties, also provides full support for it as compared with free 5-ASA. It was considered as a suitable bio-adhesive nanoparticle with mucoadhesive property to pass through the mucus layer and accumulate into the mucosa. In UC model mice, a two-fold decrease in the disease activity indexes and colon weight/length ratios was significantly observed in the group treated with 5-ASA-HbNPs. This group received one percent of the standard dosage of 5-ASA (50 μg/kg), while, a similar result was observed for a significant amount of free 5-ASA (5 mg/kg). Furthermore, microscopic images of histological sections of the extracted colons demonstrated that the 5-ASA-HbNPs and 5-ASA groups displayed instances of inflammatory damage within the colon. However, in comparison to the colitis group, the extent of this damage was relatively moderate, suggesting 5-ASA-HbNPs improved therapeutic efficacy with the lower dosage form. [Display omitted] [ABSTRACT FROM AUTHOR]

SKBR3 cells as HER2 positive cells were selectively targeted and separated with this magnetic diatom structure from the mix cells using a magnetic field. • A novel magnetic biostructures was synthesized by Chaetoceros sp. Diatoms biosilica shells. • Magnetic biosilica structure with intrinsic fluorescence was used for bioimaging. • HER2 positive cells were selectively targeted and separated by magnetic diatom structure. The ability to identify and enrich target cells can play a significant role in biosensing in general. For the separation of rare cells; a biosilica structure was extracted from " Chaetoceros sp." diatoms as a novel natural source of mesoporous materials. These diatoms had special optical capabilities, especially in fluorescence emission. Biosilica surfaces of Chaetoceros sp. were chemically modified by iron oxide nanoparticles resulting in diatom silica magnetic particles functionalized with Trastuzumab antibody to separate the breast cancer cells from normal cells. The fully characterization of magnetic biosilica structure were studied by various spectroscopic techniques. The magnetic diatom conjugated with antibody displays strong absorption and two main types of fluorescence emission with peaks centered at 493 and 650 nm (photo-excited at 405 nm). As in vitro study, SKBR3 cells (HER2 positive cells) were selectively targeted and separated with this magnetic diatom structure from the mix of HER2 negative cells using a magnetic field. These results show that Chaetoceros silica shells are promising eco-friendly biomaterials suitable for biosensing chip and the targeted delivery of drugs to the specific sites. [ABSTRACT FROM AUTHOR]

Chitosan (CHS) and chitin (CHT) biopolymers have found many applications in the field of controlled-release drug delivery systems. Herein, molecular dynamics (MD) simulation and binding free energy calculations were used to investigate the potentials of CHS and CHT polymers for the controlled release of follicle-stimulating hormone (FSH). The results indicated that FSH conformation did not change in the presence of CHS and CHT. In addition, FSH-polymer interactions caused stability of the 3 10 -helix structure of the alpha subunits of FSH (FSHα). Both the biopolymers interacted with the protein mainly through the hydrophobic forces. CHS has more affinity for FSH when compared with CHT. Furthermore, in both systems, the affinity of polymers for FSHα was more than that for beta subunits of FSH (FSHβ). The results suggested that the polysaccharides might improve the controlled-release FSH delivery. [ABSTRACT FROM AUTHOR]

Design and development of novel nanobiomaterials with multifunctional properties hold the promise of new therapeutic strategies to improve the current clinical research in the field of tissue engineering and regenerative medicine. Here, we have successfully synthesized novel autofluorescent hydroxyapatite (HAP) using a template-assisted assembly process followed by a hydrothermal method. Iron quantum cluster (FeQC) were applied as an ideal template for obtaining fluorescent HAP nanorod-assembled microspheres under different hydrothermal temperatures. We evaluated the changes in physicochemical properties, cell viability, and antibacterial activity of HAP microstructures at 120 °C, 150 °C, and 180 °C reaction temperatures. The obtained HAP samples showed an increase in size, a decrease in surface area, and also, a change in the emission wavelength range when the synthesis temperature increases. The cytotoxicity assay results of human amniotic membrane epithelial cells and C2C12 cell line revealed that the HAP with the higher specific surface area show more interaction with cells which lead to higher cytotoxicity effect. The produced HAPs displayed an inhibition zone for Escherichia coli and Staphylococcus aureus bacteria. Therefore, they showed synergistic antibacterial activity against both Gram-negative and Gram-positive bacteria. Our data clearly revealed that the presented biocompatible, multicolor, and highly stable fluorescent HAPs with considerable antibacterial and antioxidant activity have great potential for imaging applications as well as the prevention of implant-associated infections. [Display omitted] • A novel fluorescent Hydroxyapatites (HAP) were fabricated at different hydrothermal temperatures. • The fluorescent HAP samples were synthesized using iron quantum cluster as a template. • The produced HAPs exhibited multicolor fluorescence emission. • The Fluorescent HAP samples exhibited cell-type dependence cytotoxic activity. • The synthesized HAPs showed potent antibacterial effect against E.coli and S.aureus bacteria. [ABSTRACT FROM AUTHOR]

Abstract: It has been lately proposed that the interaction between like-charged residues stabilizes the native state of proteins. To explore this, we created a histidine–histidine pair in the Ca-III binding site of the Bacillus amyloliquefaciens α-amylase (BAA) and then examined the impact of this pairing on the BAA. For this purpose, we used site-directed mutagenesis (SDM) to substitute Pro407 with His, Ala, Gln, Arg, and Glu in the BAA. Subsequently, thermostability, kinetic parameters and structural properties of these variants were measured. Moreover, His–His pairing effect on the BAA thermostability was examined by simultaneous mutation of two residues (P407H/H406A and P407H/H406N). The data exhibited a significant improve in thermostability and structural features of enzyme by His replacement instead of Pro407. Other substitutions in this site did not have a significant effect on the enzyme properties, except for P407R, which yielded a partial improvement. The results also showed that the thermostabilities of double mutants significantly decreased compared with that of the P407H mutant. Moreover, the thermostability of P407H remarkably increased compared with that of other variants even in the absence of Ca2+. Our data clearly demonstrated that His406–His407 pairing was the major cause for improved thermal stability. [Copyright &y& Elsevier]

Micro-nano scale surface modification of Ti-6Al-4V was investigated through the fascinated modern fiber engraving laser method. The process was performed at a high laser speed of 2000 mm/s, under different laser frequencies (20–160 kHz) and groove distances (0.5–50 μm). Topographic evaluations such as Atomic Force Microscopy (AFM) and Field Emission Scanning Electron Microscopy (FE-SEM) were used to identify the quality and regularity of patterns. The proliferation of human osteoblast-like osteosarcoma cells (MG63) was analyzed by MTT assay for up to 72 h. Also, the plate counting method was used to quantify the viability potential of the modified surface against Escherichia coli bacteria. The cellular viability of the sample modified at the laser frequency of 20 kHz and grooving distance of 50 μm increased up to 35 and 10% compared to the non-treated and control samples, respectively. In the case of the surface modification at lower grooving distances range between 0.5–50 μm, the maximum laser frequency (160 kHz) applied leads to lower pulse's energies and less bacterial adhesion. Otherwise, at groove distances more than 50 μm, the minimum laser frequency (20 kHz) applied reduces the laser pulse overlaps, increases the cell adhesion and antibacterial properties. Surface modification by the fiber engraving laser process significantly enhances the cell adhesion on the surface. As a result of such roughness and cell adhesion enhancement, the surface toxicity feature diminished, and its antibacterial properties improved. [ABSTRACT FROM AUTHOR]

Lack of perfect insulin signaling can lead to the insulin resistance, which is the hallmark of diabetes mellitus. Activation of insulin and its binding to the receptor for signaling process initiates via B-chain C-terminal hinge conformational change through an open structure to "wide-open" conformation. Observational studies and basic scientific evidence suggest that vitamin D and E directly and/or indirectly prevent diabetes through improving glucose secretion and tolerance, activating calcium dependent endopeptidases and thus improving insulin exocytosis, antioxidant effect and reducing insulin resistance. On the contrary, clinical trials have yielded inconsistent results about the efficacy of vitamin D supplementations for the control of glucose hemostasis. In this work, best binding modes of vitamin D 3 and E on insulin obtained from AutoDock Vina were selected for Molecular Dynamic, MD, study. The binding energy obtained from Molecular Mechanics- Poisson Boltzman Surface Area, MM-PBSA method, revealed that Vitamins D 3 and E have good affinity to bind to the insulin and vitamin E has higher binding energy (−46 kj/mol) by engaging more residues in binding site. Distance and angle calculation results illustrated that vitamin E changes the B-chain conformation and it causes the formation of wide-open/active form of insulin. Vitamin E increases the ValB12-TyrB26 distance to ∼15 Å and changes the hinge angle to ∼65°. Consequently, essential hydrophobic residues for binding to insulin receptor exposed to surface in the presence of vitamin E. However, our data illustrated that vitamin D 3 cannot change B-chain conformation. Thus our MD simulations propose a model for insulin activation through vitamin E interaction for therapeutic approaches. Image 1 • Imperfection of insulin signaling can lead to insulin resistance, which is the hallmark of diabetes mellitus. • Activation of insulin and binding to its receptor initiates via B-chain C-terminal hinge conformational change. • The wide-open structure of B-chain hinge is critical for receptor binding. • Vitamin E effectively activates human insulin monomer via B-chain hinge opening. [ABSTRACT FROM AUTHOR]

Bioluminescence of a variety of marine organisms, mostly cnidarians and ctenophores, is carried out by Ca2+-dependent photoproteins. The mechanism of light emission operates via the same reaction in both animal families. Despite numerous studies on the ctenophore photoprotein family, the detailed catalytic mechanism and arrangement of amino acid residues surrounding the chromophore in this family are a mystery. Here, we report the crystal structure of Cd2+-loaded apo-mnemiopsin1, a member of the ctenophore family, at 2.15 Å resolution and used quantum mechanics/molecular mechanics (QM/MM) to investigate its reaction mechanism. The simulations suggested that an Asp-156--Arg-39--Tyr-202 triad creates a hydrogen-bonded network to facilitate the transfer of a proton from the 2-hydroperoxy group of the chromophore coelenterazine to bulk solvent. We identified a water molecule in the coelenteramide-binding cavity that forms a hydrogen bond with the amide nitrogen atom of coelenteramide, which, in turn, is hydrogen-bonded via another water molecule to Tyr-131. This observation supports the hypothesis that the function of the coelenteramide-bound water molecule is to catalyze the 2-hydroperoxycoelenterazine decarboxylation reaction by protonation of a dioxetanone anion, thereby triggering the bioluminescence reaction in the ctenophore photoprotein family. [ABSTRACT FROM AUTHOR]

Circulating miRNAs are emerging as novel reliable biomarkers for early detection of cancer diseases. Through combining the advantages of electrochemical methods and nanomaterials with the selectivity of the oligo-hybridization-based biosensors, a novel electrochemical nanobiosensor for plasma miR-155 detection have demonstrated here, based on thiolated probe-functionalized gold nanorods (GNRs) decorated on the graphene oxide (GO) sheet on the surface of the glassy carbon electrode (GCE). The reduction signals of a novel intercalating label Oracet Blue (OB), were measured by differential pulse voltammetry (DPV) method. The transmission electron microscope (TEM) imaging, UV–vis spectrophotometry, cyclic voltammetry (CV), field emission scanning electron microscope (FE-SEM) imaging and energy dispersive spectroscopy (EDS) were proved the right synthesis of the GNRs and correct assembly of the modified electrode. The electrochemical signal had a linear relationship with the concentration of the target miRNA ranging from 2.0 fM to 8.0 pM, and the detection limit was 0.6 fM. Furthermore, the nanobiosensor showed high Specificity, and was able to discriminate sharply between complementary target miRNA, single-, three-base mismatch, and non-complementary miRNA. Alongside the outstanding sensitivity and selectivity, this nanobiosensor had great storage ability, reproducibility, and showed a decent response in the real sample analysis with plasma. In conclusion, the proposed electrochemical nanobiosensor could clinically be used in the early detection of the breast cancer, by direct detection of the plasma miR-155 in real clinical samples, without a need for sample preparation, RNA extraction and/or amplification. [ABSTRACT FROM AUTHOR]

[Display omitted] • A new strategy was developed to deliver 5-ASA to an inflamed site in the colon by HbNPs. • This conjugated drug is protected against chemical and enzymatic hydrolysis. • Higher cellular uptake and prolonged release time for 5-ASA-HbNPs in the Caco-2 cells. • HbNPs increase the effectiveness of drug therapy due to adhesion to intestinal epithelial cells. • In vivo imaging of mouse colon shows the localization of drug in the descending colon. A colonic drug delivery system was developed to specifically deliver 5-aminosalicylic acid (5-ASA) to the inflamed site by conjugating with hemoglobin nanoparticles (HbNPs). The 5-ASA-HbNPs (eight 5-ASA molecules per Hb molecule) with the size of 220 nm and zeta potential of −14.6 mV is a tailored nanoparticle able to pass through the mucus layer. The 5-ASA-HbNPs do not undergo chemical and enzymatic hydrolysis in the simulated gastrointestinal fluids over 6 h. Significantly higher cellular uptakes and prolonged release was seen for the 5-ASA-HbNPs in Caco-2 cells, compared to free 5-ASA over 72 h. In addition, 5-ASA-HbNPs revealed similar therapeutic effectiveness with free 5-ASA against tumor necrosis factor and showed less inhibitory concentration (IC50) for myeloperoxidase enzyme activity. In vivo imaging of mouse demonstrated the localization of drug in the descending colon after oral administration and about 15% of the administered dose was recovered as 5-ASA from urine in 6 h. The use of these nanoparticles with the mucus adhesion properties and permeability to intestinal epithelial cells can be a good candidate with potential application in the colonic drug delivery field. [ABSTRACT FROM AUTHOR]

Abstract: Some new phosphorus(V) hydrazides 1a–12a were synthesized and characterized by 1H, 13C, 31P NMR, IR spectroscopy and elemental analysis. Moreover, the interaction of Cu(M)2·nH2O with 1a, 3a and 7a gave 4,4′-bis(morpholine)diazene (1b). In fact, in these reactions, copper(II) ions acted as oxidizing agent. The results supported the proposed mechanism. The structures of compounds 1a, 1b and 1c were further determined by X-ray crystallography. Compounds 1a–12a were screened for their antibacterial activities. Also, the acetyl- and butyrylcholinesterase inhibitory activity of 1a, 3a, 7a, 11a and 12a was measured using Ellman’s method. It is interesting that these compounds were more potent inhibitors of BChE than of AChE. Also, using Lineweaver–Burk plots, it was indicated these compounds are mixed inhibitors. [ABSTRACT FROM AUTHOR]

Abstract: A facile method for the formation of covalent bonds between protein molecules is zero-length cross-linking. This method enables the formation of cross-links without use of any chemical reagents. Here, the cross-linking is performed for lysozyme, peroxidase (a glycoprotein) and between lysozyme–peroxidase by the method of Simons et al. [B.L. Simons, M.C. King, T. Cyr, M.A. Hefford, H. Kaplan, Covalent cross-linking of protein without chemical reagents, Protein Sci. 2002, 11, 1558–1564]. Approximately one-third of the total lysozyme becomes cross-linked and the dimer form was the major product for both enzymes. This modification induced some changes in the kinetic properties of the dimer peroxidase, as evident by two-fold increasing of V max compared to the monomer but the enzymatic activity of cross-linked lysozyme dimer was the same as monomer. The activity of lysozyme dimer remained constant up to 10min at 80°C, while peroxidase activity of both monomer and dimer began to decrease after heating. The structural changes of the enzymes were investigated by circular dichroism and intrinsic fluorescence techniques. Near UV result showed lysozyme possess a compact structure in the dimer form but disruption of tertiary structure of peroxidase dimer was observed. Also conformational changes were detected and discussed by intrinsic fluorescence experiments. Effect of several metals in the formation of lysozyme dimer showed that Co2+ is the most effective one but its effect was marginal. At the end formation of heterogeneous dimer, peroxidase–lysozyme, was achieved using this method. [Copyright &y& Elsevier]

Abstract: A native α-amylase capable of activity at low pH was isolated from Bacillus sp. KR-8104 (KRA) naturally occurred in rhizoplane zone of potato cultured soils. An investigation was set out to account some evidence for its specific properties. Kinetic parameters of KRA with respect to Bacillus licheniformis α-amylase (BLA) and B. amyloliquefaciens α-amylase (BAA) showed a shift in acidic limb for KRA pH-profile. After isolation of KRA gene and determination of its nucleotide and protein sequences, the three-dimensional structure of KRA was simulated based on BLA as template. Using PROPKA program the pK a values of titrable groups in KRA model together with other few members of this family were predicted either from simulated model or available structure at protein data bank. A correlation between catalytic residue pK a values and the optimal pH-activity profile was seen. Both our experimental data and prediction studies indicated a shift in pK a value of catalytic residues to acidic pH. Moreover alignment studies raised amino acid substitutions in KRA compare to BLA which may affect the KRA putative active site leading to the formation of an extra hydrogen bond between Glu261 and Arg229 (regarding to BLA numbering) and in turn shifts the pH-activity profile to lower amount. Also a common intermediate with characteristics of molten globular state was detected at a pH value lower than other members of α-amylase family using far UV circular dichroism, intrinsic and ANS fluorescence and quenching experiments. [Copyright &y& Elsevier]

The firefly bioluminescence reaction, which uses luciferin, Mg-ATP, and molecular oxygen to yield an electronically excited oxyluciferin, is carried out by luciferase and visible light is emitted. The bioluminescence color of firefly luciferases is determined by the luciferase structure and assay conditions. Among different beetle luciferases, those from Phrixothrix railroad worm emit either yellow or red bioluminescence colors. Sequence alignment analysis shows that the red-emitter luciferase from Phrixothrix hirtus has an additional Arg residue at 353, which is absent in firefly luciferases. We report here the construction and purification of a mutant at residue Arg356, which is not conserved in beetle luciferases. By insertion of an additional residue (Arg356) using site-specific insertion mutagenesis in a green-emitter luciferase (Lampyris turkestanicus) the color of emitted light was changed to red and the optimum temperature of activity was also increased. Insertion of this Arg in an important flexible loop showed changes of the bioluminescence color and the luciferase reaction took place with relatively retention of its basic kinetic properties such as Km and relative activity. Comparison of native and mutant luciferases using homology modeling reveals a significant conformational change of the flexible loop in the red mutant. Movement of flexible loop brought about a new ionic interaction concomitant with a change in polarity of the emitter site, thereby leading to red emission. It is worthwhile to note that the increased optimum temperature and emission of red light might make mutant luciferase a suitable reporter for the study of gene expression and bioluminescence imaging. [ABSTRACT FROM AUTHOR]

Abstract: Bacterial polyphenol oxidases (PPOs) with a high oxidizing capacity for a wide range of substrates could make their applications in phenolic biotransformation, food processing, cosmetics, and textile industry. We have isolated a melanogenic soil bacterium by differential screening of a number of strains which were isolated from the Iranian microflora. The taxonomic characterization of this strain indicates that it belongs to the genus Bacillus (HR03), and has the ability to produce all types of PPOs; cresolase (EC 1.14.18.1), cathecolase (EC.1.10.3.1), and laccase (EC 1.10.3.2). We studied the tyrosinase activity using l-tyrosine and l-dopa as substrates and the laccase activity with specific substrates such as syringaldazine and 2,6-dimethoxyphenol. The optimum pH and temperature, obtained for all types of polyphenol oxidases, were at about pH 5.5 and 55°C, respectively. Tyrosinase-like enzyme of this strain shows a lag period in its tyrosine hydroxylase activity that could be avoided by the addition of small amounts of l-dopa and sodium dodecyl sulfate (SDS). In addition, tyrosinase and laccase were activated by SDS below the critical micelle concentration and were inhibited by 1mM EDTA. We tested the resistance of melanized-cells against UVA, UVC and H2O2. Results show that melanin protects strain HR03 against UV lights and the oxidant. [Copyright &y& Elsevier]

Abstract: In this study, we report for the first time, production of monoclonal antibody (MAb) against beta human chorionic gonadotropin (hCG) with proteolytic activity. MAb “7D9” was raised in Balb/C mice using purified human chorionic gonadotropin. Immunoblot analysis and enzyme-linked immunoabsorbent assay (ELISA) showed that this MAb reacts with beta hCG. The epitope for this antibody appears to be located in the C-terminal of beta chain as suggested by the absence of cross-reaction with other glycoprotein hormones such as FSH, TSH and LH. Our data reveal that this MAb is very unstable and has autodegradation characteristics. Zymogram analyses also show that 7D9 MAb has a high level of hydrolytic activity against different substrates such as casein and gelatin. This proteolytic activity can be inhibited by EDTA. These findings demonstrate the proteolytic character of 7D9 MAb and consequently explain its instability. [Copyright &y& Elsevier]

Abstract: An investigation was carried out to compare the proteolytic resistance and acidic digestion of the mesophilic α-amylase from Bacillus amyloliquefaciens (BAA) and its thermophilic counterpart from Bacillus licheniformis (BLA). Correlation between sites of proteolytic cleavage and the three-dimensional structure of the α-amylases, with the application of theoretical modeling of BAA, allowed discussion of the flexibility and the stability of both enzymes. The thermophilic enzyme shows higher resistance to trypsin, papain and thermolysin but is sensitive to pronase and acidic digestion. Proteolytic digestion of the thermophilic enzyme leads to an increased activity of the enzyme at room temperature whereas results of SDS-PAGE indicate proteolytic cleavage. Furthermore, thermal stability and resistance to proteolysis for BLA and BAA in the presence of additives such as sorbitol, trehalose and glycerol were also investigated. In addition to thermal stabilization of the two enzymes, these additives also augmented the resistance of the enzymes to proteolysis. [Copyright &y& Elsevier]