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Protein Matrix (protein + matrix)
Selected AbstractsHigh-Resolution Patterning of Hydrogels in Three Dimensions using Direct-Write Photofabrication for Cell GuidanceADVANCED FUNCTIONAL MATERIALS, Issue 22 2009Stephanie K. Seidlits Abstract The development of three-dimensional, spatially defined neuronal cultures that mimic chemical and physical attributes of native tissue is of considerable interest for various applications, including the development of tailored neuronal networks and clinical repair of damaged nerves. Here, the use of multiphoton excitation to photocrosslink protein microstructures within three-dimensional, optically transparent hydrogel materials, such as those based on hyaluronic acid, is reported. Multiphoton excitation confines photocrosslinking to a three-dimensional voxel with submicron spatial resolution, enabling fabrication of protein matrices with low- to sub-micrometer feature sizes by scanning the focus of a laser relative to the reagent solution. These methods can be used to create complex three-dimensional architectures that provide both chemical and topographical cues for cell culture and guidance, providing for the first time a means to direct cell adhesion and migration on size scales relevant to in vivo environments. Using this approach, guidance of both dorsal root ganglion cells (DRGs) and hippocampal neural progenitor cells (NPCs) along arbitrary, three-dimensional paths is demonstrated. [source] Morphological and biochemical analyses of otoliths of the ice-fish Chionodraco hamatus confirm a common origin with red-blooded speciesJOURNAL OF ANATOMY, Issue 1 2009Chiara Maria Motta Abstract The morphology and composition of the three otoliths of the Antarctic ice-fish Chionodraco hamatus were studied by scanning electron microscopy and X-ray diffraction. The composition of the sagitta, lapillus and asteriscus protein matrices was also analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, western blots and confocal laser scanning microscopy to reveal the presence of and to localize the calcium-binding proteins calmodulin, calbindin and S-100. Morphological results indicated that the otoliths in this ice-fish were similar to those of Trematomus bernacchii, a red-blooded Antarctic species [B. Avallone et al. (2003) J. Submicrosc. Cytol. Pathol. 35, 69,76], but rather different from those of other teleosts. These two Antarctic species possessed a completely vateritic asteriscus, whereas their sagitta and lapillus were made mostly of aragonite. Parallel analysis of protein patterns in C. hamatus and T. bernacchii revealed that the sagitta significantly differed from the lapillus and asteriscus in both species. The sagitta did not contain the S-100 protein and showed calmodulin and calbindin located in discontinuous or incremental zones, respectively. These results demonstrate that the otoliths of C. hamatus and T. bernacchii share more resemblances than differences and support the idea of a common origin of these species. [source] RHEOLOGY AND MICROSTRUCTURE OF WHEAT DOUGH DEVELOPED WITH CONTROLLED DEFORMATIONJOURNAL OF TEXTURE STUDIES, Issue 1 2000EMILY J. SCHLUENTZ ABSTRACT Undeveloped wheat dough samples were strained in shear and extensional flow between parallel plates to produce a controlled level of development. Dough made in a standard Farinograph, considered developed dough, was used for comparison. Scanning electron microscopy images of deformed dough were subjected to numerical image processing to characterize the protein matrix present. Results were compared to dynamic rheological properties to evaluate the influence of strain deformation on the formation of microstructure. Viscoelastic moduli of wheat dough showed that developed dough had the greatest amount of structure formation, followed by extensionally-strained and shear-strained samples, respectively. Undeveloped dough showed the lowest levels of structure development. Image analysis indicated statistically significant differences between protein matrices in developed and undeveloped samples; however, results were not significantly different between shear- and extension-ally-strained samples. [source] Probing protein structure and dynamics by second-derivative ultraviolet absorption analysis of cation,, interactionsPROTEIN SCIENCE, Issue 10 2006Laura H. Lucas Abstract We describe an alternate approach for studying protein structure using the detection of ultraviolet (UV) absorbance peak shifts of aromatic amino acid side chains induced by the presence of salts. The method is based on the hypothesis that salt cations (Li+, Na+, and Cs+) of varying sizes can differentially diffuse through protein matrices and interact with benzyl, phenyl, and indole groups through cation,, interactions. We have investigated the potential of this method to probe protein dynamics by measuring high resolution second-derivative UV spectra as a function of salt concentration for eight proteins of varying physical and chemical properties and the N -acetylated C -ethyl esterified amino acids to represent totally exposed side chains. We show that small shifts in the wavelength maxima for Phe, Tyr, and Trp in the presence of high salt concentrations can be reliably measured and that the magnitude and direction of the peak shifts are influenced by several factors, including protein size, charge, and the local environment and solvent accessibility of the aromatic groups. Evaluating the empirical UV spectral data in light of known protein structural information shows that probing cation,, interactions in proteins reveals unique information about the influence of structure on aromatic side chain spectroscopic behavior. [source] Functionalized-Silk-Based Active Optofluidic DevicesADVANCED FUNCTIONAL MATERIALS, Issue 7 2010Konstantinos Tsioris Abstract Silk protein from the silkworm Bombyx mori has excellent chemical and mechanical stability, biocompatibility, and optical properties. Additionally, when the protein is purified and reformed into materials, the biochemical functions of dopants entrained in the protein matrix are stabilized and retained. This unique combination of properties make silk a useful multifunctional material platform for the development of sensor devices. An approach to increase the functions of silk-based devices through chemical modifications to demonstrate an active optofluidic device to sense pH is presented. Silk protein is chemically modified with 4-aminobenzoic acid to add spectral-color-responsive pH sensitivity. The functionalized silk is combined with the elastomer poly(dimethyl siloxane) in a single microfluidic device. The microfluidic device allows spatial and temporal control of the delivery of analytic solutions to the system to provide the optical response of the optofluidic device. The modified silk is stable and spectrally responsive over a wide pH range from alkaline to acidic. [source] Use of a ,-glucan hydrocolloidal suspension in the manufacture of low-fat Cheddar cheese: manufacture, composition, yield and microstructureINTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 1 2004G. Konuklar Summary Low-fat Cheddar cheese was manufactured using a , -glucan, hydrocolloidal fat replacer denoted as Nutrim. The composition, production efficiency, microstructure, and utility of replacing fat with Nutrim were examined. Cheese samples (designated as Nutrim-I, and Nutrim-II) containing Nutrim were produced with mean fat levels of 6.84 and 3.47%, respectively. A low-fat cheese was also produced as a control with a mean fat level of 11.2%. Nutrim-II cheese had significantly higher moisture, salt, and ash contents as compared with the low-fat control cheese. The low-fat control cheese had a higher yield normalized for 54% moisture and 1.5% salt content as compared with the Nutrim-II cheese. Scanning electron microscopy revealed smaller and more uniform fat droplet voids in the Nutrim cheese than the low-fat control, and a more dense, noncontinuous background protein matrix with globular clusters suggesting a physical buffering afforded by the presence of the , -glucan hydrocolloid or its associated water. [source] [Fe-Fe]-hydrogenase reactivated by residue mutations as bridging carbonyl rearranges: A QM/MM studyINTERNATIONAL JOURNAL OF QUANTUM CHEMISTRY, Issue 14 2010Stefan Motiu Abstract In this work, we found aqueous enzyme phase reaction pathways for the reactivation of the exogenously inhibited [Fe-Fe]-hydrogenases by O2, or OH,, which metabolizes to H2O (Dogaru et al., Int J Quantum Chem 2008, 108; Motiu et al., Int J Quantum Chem 2007, 107, 1248). We used the hybrid quantum mechanics/molecular mechanics (QM/MM) method to study the reactivation pathways of the exogenously inhibited enzyme matrix. The ONIOM calculations performed on the enzyme agree with experimental results (Liu et al., J Am Chem Soc 2002, 124, 5175), that is, wild-type [Fe-Fe]-hydrogenase H-cluster is inhibited by oxygen metabolites. An enzyme spherical region with a radius of 8 Å (from the distal iron, Fed) has been screened for residues that prevent H2O from leaving the catalytic site and reactivate the [Fe-Fe]-hydrogenase H-cluster. In the screening process, polar residues were removed, one at a time, and frequency calculations provided the change in the Gibbs' energy for the dissociation of water (due to their deletion). When residue deletion resulted in significant Gibbs' energy decrease, further residue substitutions have been carried out. Following each substitution, geometry optimization and frequency calculations have been performed to assess the change in the Gibbs' energy for the elimination of H2O. Favorable thermodynamic results have been obtained for both single residue removal (,G,Glu374 = ,1.6 kcal/mol), single substitution (,GGlu374His = ,3.1 kcal/mol), and combined residue substitutions (,GArg111Glu;Thr145Val;Glu374His;Tyr375Phe = ,7.5 kcal/mol). Because the wild-type enzyme has only an endergonic step to overcome, that is, for H2O removal, by eliminating several residues, one at a time, the endergonic step was made to proceed spontaneously. Thus, the most promising residue deletions which enhance H2O elimination are ,Arg111, ,Thr145, ,Ser177, ,Glu240, ,Glu374, and ,Tyr375. The thermodynamics and electronic structure analyses show that the bridging carbonyl (COb) of the H-cluster plays a concomitant role in the enzyme inhibition/reactivation. In gas phase, COb shifts towards Fed to compensate for the electron density donated to oxygen upon the elimination of H2O. However, this is not possible in the wild-type enzyme because the protein matrix hinders the displacement of COb towards Fed, which leads to enzyme inhibition. Nevertheless, enzyme reactivation can be achieved by means of appropriate amino acid substitutions. © 2009 Wiley Periodicals, Inc. Int J Quantum Chem, 2010 [source] Embryonic undifferentiated cells show scattering activity on a surface coated with immobilized E-cadherinJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2008Masato Nagaoka Abstract Rearrangement of cell,cell adhesion is a critical event in embryonic development and tissue formation. We investigated the regulatory function of E-cadherin, a key adhesion protein, in the developmental process by using E-cadherin/IgG Fc fusion protein as an adhesion matrix in cell culture. F9 embryonal carcinoma cells usually form colonies when cultured on gelatin or fibronectin matrices. However, F9 cells cultured on the E-cadherin/IgG Fc fusion protein matrix formed a scattered distribution, with a different cytoskeletal organization and E-cadherin-rich protrusions that were regulated by Rac1 activity. The same scattering activity was observed in P19 embryonal carcinoma cells. In contrast, three types of differentiated cells, NMuMG mammary gland cells, MDCK kidney epithelial cells, and mouse primary isolated hepatocytes, did not show the scattering activity observed in F9 and P19 cells. These results suggest that migratory behavior on an E-cadherin-immobilized surface is only observed in embryonic cells, and that the regulatory mechanisms underlying E-cadherin-mediated cell adhesion vary with the state of differentiation. J. Cell. Biochem. 103: 296,310, 2008. © 2007 Wiley-Liss, Inc. [source] Interactions of ,-carrageenan Plus Other Hydrocolloids in Fish Myosystem GelsJOURNAL OF FOOD SCIENCE, Issue 6 2001M. Pérez-Mateos ABSTRACT: Mixtures of ,-carrageenan plus other hydrocolloids (locust bean, guar, xanthan, iota-carrageenan, sodium carboxymethylcellulose, and sodium alginate) were examined for their effects on the mechanical and water holding properties of heat-induced gels made from washed blue whiting mince. Gel structure and thermal behavior were also studied. No synergistic effect was detectable through functional properties except for the mixture of ,-carrageenan with locust bean gum. Light microscopy revealed that ,-carrageenan and xanthan mixed locally with locust bean at its rich domains. ,-carrageenan and xanthan presented interactions with the protein matrix, which were more discernible in the first case. Differential scanning calorimetry (DSC) revealed faint interactions for the mixtures of ,-carrageenan with locust bean and with xanthan, and weakly synergistic gelling effects between the last two hydrocolloids. The blend of ,-carrageenan with sodium alginate exhibited thermally strong synergistic interactions but no particular effects were induced on corresponding functional properties. [source] INFLUENCE OF HYDROXYPROPYL METHYLCELLULOSE ON THE RHEOLOGICAL AND MICROSTRUCTURAL CHARACTERISTICS OF WHOLE WHEAT FLOUR DOUGH AND QUALITY OF PURIJOURNAL OF TEXTURE STUDIES, Issue 2 2009M. L. SUDHA ABSTRACT Puri is a traditional unleavened fried product prepared from whole wheat flour. Hydroxypropyl methylcellulose (HPMC) was used to study its effect on rheological characteristics of whole wheat flour dough and puri making quality. Addition of HPMC at 0.5 and 1.0% w/w increased the water absorption and dough stability whereas the resistance to extension and extensibility decreased. Pasting temperature, peak viscosity and cold paste viscosity gradually decreased. The moisture and fat contents of puri increased marginally. Quality parameters and sensory acceptability were monitored after 0 and 8 h of storage. Addition of 0.5% HPMC gave higher sensory scores. Microscopic observations during puri processing showed that the starch granules in the control dough were clearly visible in the protein matrix, which reduced on frying due to partial gelatinization. Microstructure of puri with HPMC showed higher gelatinization of starch. It also helped in moisture retention and hence, resulted in highly pliable and soft-textured puri. PRACTICAL APPLICATIONS Puri is a traditional unleavened fried product that is prepared by mixing whole wheat flour and water, sheeted to a desirable thickness and fried. Use of hydroxypropyl methylcellulose (HPMC) affected the whole wheat flour dough and puri making quality. It helped in moisture retention and hence, resulted in highly pliable and soft-textured puri. Microstructure of puri with HPMC showed higher gelatinization of starch. [source] Circular Dichroism of the Photoreceptor Pigment OxyblepharisminPHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 6 2005Osvaldo Pieroni ABSTRACT Circular dichroism (CD) was used to study the structure of oxyblepharismin (OxyBP), the photoreceptor chromophore for the photophobic response of the blue form of Blepharisma japonicum. Both the chromophore associated to its native protein and the free chromophore in ethanol solution were investigated. CD spectra in the far-UV range indicate that OxyBP induces a slight increase in the ,-helix content of the protein matrix. CD spectra in the near-UV and visible region of the spectrum show that OxyBP adopts a chiral conformation with a preferential geometry not only when associated to its protein matrix, but also when isolated and dissolved in ethanol. This experimental result is related to the existence of a high-energy interconversion barrier between two enantiomeric structures of the molecule and discussed on the basis of an asymmetric biosynthesis of its precursor, blepharismin. [source] Protein Matrix Elasticity Determined by Fluorescence Anisotropy of Its Tryptophan Residues,PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 1 2003Christian Zentz ABSTRACT Rotational motions of Trp residues embedded within human hemoglobin matrix have been measured by using their steady-state fluorescence anisotropy. The mean square angular displacement ,2 of Trp residues, depending on the temperature, can be expressed by where W is the thermal energy acting on the Trp residues and C the resilient torque constant of the protein matrix. To study the external medium influencing the protein dynamics, comparative experiments were made with protein in aqueous buffer and in the presence of 32% glycerol. The data show that between 5°C and 25°C, external medium acts on the protein matrix elasticity. [source] Tracking ligand-migration pathways of carbonmonoxy myoglobin in crystals at cryogenic temperaturesACTA CRYSTALLOGRAPHICA SECTION A, Issue 2 2010Ayana Tomita In order to explore the ligand-migration dynamics in myoglobin induced by photodissociation, cryogenic X-ray crystallographic investigations of carbonmonoxy myoglobin crystals illuminated by continuous wave and pulsed lasers at 1,15,kHz repetition rate have been carried out. Here it is shown that this novel method, extended pulsed-laser pumping of carbonmonoxy myoglobin, promotes ligand migration in the protein matrix by crossing the glass transition temperature repeatedly, and enables the visualization of the migration pathway of the photodissociated ligands in native Mb at cryogenic temperatures. It has revealed that the migration of the CO molecule into each cavity induces structural changes of the amino-acid residues around the cavity which result in the expansion of the cavity. The sequential motion of the ligand and the cavity suggests a self-opening mechanism of the ligand-migration channel arising by induced fit. [source] Seed-specific expression of the wheat puroindoline genes improves maize wet milling yieldsPLANT BIOTECHNOLOGY JOURNAL, Issue 8 2009Jinrui Zhang Summary The texture of maize (Zea mays L.) seeds is important to seed processing properties, and soft dent maize is preferred for both wet-milling and livestock feed applications. The puroindoline genes (Pina and Pinb) are the functional components of the wheat (Triticum aestivum L.) Hardness locus and together function to create soft grain texture in wheat. The PINs (PINA and PINB) are believed to act by binding to lipids on the surface of starch granules, preventing tight adhesion between starch granules and the surrounding protein matrix during seed maturation. Here, maize kernel structure and wet milling properties were successfully modified by the endosperm-specific expression of wheat Pins (Pina and Pinb). Pins were introduced into maize under the control of a maize ,- Zein promoter. Three Pina/Pinb expression positive transgenic lines were evaluated over two growing seasons. Textural analysis of the maize seeds indicated that the expression of PINs decreased adhesion between starch and protein matrix and reduced maize grain hardness significantly. Reduction in pressure required to fracture kernels ranged from 15.65% to 36.86% compared with control seeds. Further, the PINs transgenic maize seeds had increased levels of extractable starch as characterized by a small scale wet milling method. Starch yield was increased by 4.86% on average without negatively impacting starch purity. The development of softer maize hybrids with higher starch extractability would be of value to maize processors. [source] Protein dynamics control proton transfer from bulk solvent to protein interior: A case study with a green fluorescent proteinPROTEIN SCIENCE, Issue 7 2005Anoop M. Saxena Abstract The kinetics of proton transfer in Green Fluorescent Protein (GFP) have been studied as a model system for characterizing the correlation between dynamics and function of proteins in general. The kinetics in EGFP (a variant of GFP) were monitored by using a laser-induced pH jump method. The pH was jumped from 8 to 5 by nanosecond flash photolysis of the "caged proton," o -nitrobenzaldehyde, and subsequent proton transfer was monitored by following the decrease in fluorescence intensity. The modulation of proton transfer kinetics by external perturbants such as viscosity, pH, and subdenaturing concentrations of GdnHCl as well as of salts was studied. The rate of proton transfer was inversely proportional to solvent viscosity, suggesting that the rate-limiting step is the transfer of protons through the protein matrix. The rate is accelerated at lower pH values, and measurements of the fluorescence properties of tryptophan 57 suggest that the enhancement in rate is associated with an enhancement in protein dynamics. The rate of proton transfer is nearly independent of temperature, unlike the rate of the reverse process. When the stability of the protein was either decreased or increased by the addition of co-solutes, including the salts KCl, KNO3, and K2SO4, a significant decrease in the rate of proton transfer was observed in all cases. The lack of correlation between the rate of proton transfer and the stability of the protein suggests that the structure is tuned to ensure maximum efficiency of the dynamics that control the proton transfer function of the protein. [source] Generation of high-quality protein extracts from formalin-fixed, paraffin-embedded tissuesPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 15 2009Maria Filippa Addis Abstract A wealth of information on proteins involved in many aspects of disease is encased within formalin-fixed paraffin-embedded (FFPE) tissue repositories stored in hospitals worldwide. Recently, access to this "hidden treasure" is being actively pursued by the application of two main extraction strategies: digestion of the entangled protein matrix with generation of tryptic peptides, or decrosslinking and extraction of full-length proteins. Here, we describe an optimised method for extraction of full-length proteins from FFPE tissues. This method builds on the classical "antigen retrieval" technique used for immunohistochemistry, and allows generation of protein extracts with elevated and reproducible yields. In model animal tissues, average yields of 16.3,,g and 86.8,,g of proteins were obtained per 80,mm2 tissue slice of formalin-fixed paraffin-embedded skeletal muscle and liver, respectively. Protein extracts generated with this method can be used for the reproducible investigation of the proteome with a wide array of techniques. The results obtained by SDS-PAGE, western immunoblotting, protein arrays, ELISA, and, most importantly, nanoHPLC-nanoESI-Q-TOF MS of FFPE proteins resolved by SDS-PAGE, are presented and discussed. An evaluation of the extent of modifications introduced on proteins by formalin fixation and crosslink reversal, and their impact on quality of MS results, is also reported. [source] Biglycan Overexpression on Tooth Enamel Formation in Transgenic MiceTHE ANATOMICAL RECORD : ADVANCES IN INTEGRATIVE ANATOMY AND EVOLUTIONARY BIOLOGY, Issue 10 2008Xin Wen Abstract Previously, it was shown that the volume of forming enamel of molar teeth in biglycan-null mice was greater than that in genetically matched wild-type mice. This phenotypic change appeared to result from an increase in amelogenin expression, implying that biglycan directly influences amelogenin synthesis. To determine whether biglycan overexpression resulted in decreased amelogenin expression, we engineered transgenic mice to overexpress biglycan in the enamel organ epithelium. Biglycan overexpression did not significantly affect the amelogenin expression in incisor and molar teeth in 3-day postnatal transgenic mice. In the transgenic animals, we observed that the immature and mature enamel appeared normal. These results suggested that increasing the biglycan expression, in the cells that synthesize the precursor protein matrix for enamel, has a negligible influence on amelogenesis. Anat Rec, 2008. © 2008 Wiley-Liss, Inc. [source] Molecular chemical structure of barley proteins revealed by ultra-spatially resolved synchrotron light sourced FTIR microspectroscopy: Comparison of barley varietiesBIOPOLYMERS, Issue 4 2007Peiqiang Yu Abstract Barley protein structure affects the barley quality, fermentation, and degradation behavior in both humans and animals among other factors such as protein matrix. Publications show various biological differences among barley varieties such as Valier and Harrington, which have significantly different degradation behaviors. The objectives of this study were to reveal the molecular structure of barley protein, comparing various varieties (Dolly, Valier, Harrington, LP955, AC Metcalfe, and Sisler), and quantify protein structure profiles using Gaussian and Lorentzian methods of multi-component peak modeling by using the ultra-spatially resolved synchrotron light sourced Fourier transform infrared microspectroscopy (SFTIRM). The items of the protein molecular structure revealed included protein structure ,-helices, ,-sheets, and others such as ,-turns and random coils. The experiment was performed at the National Synchrotron Light Source in Brookhaven National Laboratory (BNL, US Department of Energy, NY). The results showed that with the SFTIRM, the molecular structure of barley protein could be revealed. Barley protein structures exhibited significant differences among the varieties in terms of proportion and ratio of model-fitted ,-helices, ,-sheets, and others. By using multi-component peaks modeling at protein amide I region of 1710,1576 cm,1, the results show that barley protein consisted of approximately 18,34% of ,-helices, 14,25% of ,-sheets, and 44,69% others. AC Metcalfe, Sisler, and LP955 consisted of higher (P < 0.05) proportions of ,-helices (30,34%) than Dolly and Valier (,-helices 18,23%). Harrington was in between which was 25%. For protein ,-sheets, AC Metcalfe, and LP955 consisted of higher proportions (22,25%) than Dolly and Valier (13,17%). Different barley varieties contained different ,-helix to ,-sheet ratios, ranging from 1.4 to 2.0, although the difference were insignificant (P > 0.05). The ratio of ,-helices to others (0.3 to 1.0, P < 0.05) and that of ,-sheets to others (0.2 to 0.8, P < 0.05) were different among the barley varieties. It needs to be pointed out that using a multi-peak modeling for protein structure analysis is only for making relative estimates and not exact determinations and only for the comparison purpose between varieties. The principal component analysis showed that protein amide I Fourier self-deconvolution spectra were different among the barley varieties, indicating that protein internal molecular structure differed. The above results demonstrate the potential of the SFTIRM to localize relatively pure protein areas in barley tissues and reveal protein molecular structure. The results indicated relative differences in protein structures among the barley varieties, which may partly explain the biological differences among the barley varieties. Further study is needed to understand the relationship between barley molecular chemical structure and biological features in terms of nutrient availability and digestive behavior. © 2006 Wiley Periodicals, Inc. Biopolymers 85:308,317, 2007. This article was originally published online as an accepted preprint. The "Published Online" date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com [source] A method for lipase co-precipitation in a biodegradable protein matrixBIOTECHNOLOGY & BIOENGINEERING, Issue 6 2007M. Golubovic Abstract This article presents a novel method for immobilization of active ingredients. The method is based on CO2 aided active ingredient co-precipitation with glycinin, a biodegradable protein matrix from edible soybean protein. Glycinin precipitates abundantly under isoelectric conditions and serves as the matrix within which the active substance is trapped during the precipitation process. The enzyme lipase from Candida rugosa was successfully co-precipitated into the protein pellet to prove the principle. It was shown that the lipase within the co-precipitate retained lipase and esterase activity under different pH conditions. In some cases the activity was even higher than the activity of crude lipase, possibly due to the protective role of the matrix protein. Due to the retained lipase activity and food-grade quality of the binary precipitate, it has potential of being used in the food or pharmaceutical industry. Additional quality of the binary precipitate is the potentially significantly reduced downstream processing due to the fact that no organic solvents or precipitants were used in the precipitation process. Biotechnol. Bioeng. 2007;98: 1209,1218. © 2007 Wiley Periodicals, Inc. [source] Systematic Regulation of the Enzymatic Activity of Phenylacetaldoxime Dehydratase by Exogenous LigandsCHEMBIOCHEM, Issue 12 2006Katsuaki Kobayashi Dr. Abstract Phenylacetaldoxime dehydratase from Bacillus sp. OxB-1 (OxdB) contains a heme that acts as the active site for the dehydration reaction of aldoxime. Ferrous heme is the active form, in which the heme is five coordinate with His282 as a proximal ligand. In this work, we evaluated the functional role of the proximal ligand for the catalytic properties of the enzyme by "the cavity mutant technique". The H282G mutant of OxdB lost enzymatic activity, although the heme, which was five coordinate with a water molecule (or OH,) as an axial ligand, existed in the protein matrix. The enzymatic activity was rescued by imidazole or pyridine derivatives that acted as the exogenous proximal ligand. By changing the electron-donation ability of the exogenous ligand with different substituents, the enzymatic activity could be regulated systematically. The stronger the electron-donation ability of the exogenous ligand, the higher was the restored enzymatic activity. Interestingly, H282G OxdB with 2-methyl imidazole showed a higher activity than the wild-type enzyme. Kinetic analyses revealed that the proximal His regulated not only the affinity of substrate binding to the heme but also the elimination of the OH group from the substrate. [source] Human Fallopian Tube Neutrophils , A Distinct Phenotype from Blood NeutrophilsAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 4 2006Jennifer M. Smith Problem, The role of neutrophils in the human Fallopian tube (FT) is unknown. In order to provide insights into their functions in the FT, we systematically compared neutrophils from normal FT and peripheral blood (PB). Method of study, Flow cytometric analysis of surface receptors, granule proteins, and intracellular cytokines expressed by neutrophils from enzymatically dispersed FT and PB was performed. Results, Fallopian tube neutrophils expressed significantly higher levels of CD64, human class II histocompatibility antigen DR (HLA-DR), , -interferon, and vascular endothelial growth factor than those from PB. Fewer FT neutrophils expressed IL-8 receptors compared to PB, while more expressed the receptor for the bacterial-derived chemoattractant formyl-Met-Leu-Phe (fMLP). The number of FT neutrophils containing the granule proteins matrix metalloproteinase-9, lactoferrin, and myeloperoxidase was decreased versus PB. Conclusion, Fallopian tube neutrophils exhibit a phenotype distinct from PB neutrophils, suggesting functional activation of innate immune defense in the female reproductive tract as well as a potential role in maintaining normal FT physiology. [source] | |||||||||||||||||||||||||