Gel Electrophoretic Patterns Research Articles

Genetic relationship study of some Vicia species by FISH and total seed storage protein patterns

BackgroundGenus Vicia is a member of family Fabaceae and comprises 180 to 210 species. The most important species is faba bean (Vicia faba) which is still one of the most favourable grain legumes over all the world. The genus contains some additional food crops and a number of forage plants and some other weedy strains such as Vicia angustifolia and Vicia cordata. The aim of the present investigation is to elucidate the phylogenetic relationships among four Vicia species, two species (Vicia angustifolia L. ssp. Angustifolia (2n = 12) and Vicia cordata wulfen ex Hoppe (2n = 10)) belong to section Vicia, Vicia dalmatica A. Kern (2n = 12, section Cracca), and Vicia johannis tamamsch (2n = 14, section Faba). ResultsTwo tools have been applied to identify the genetic relationships among the examined species, double fluorescence in situ hybridization (FISH) has been used to localize the sites of 5S and 45S rDNA, and sodium dodecyl sulfate-poly acrylamide gel electrophoretic (SDS-PAGE) patterns of total seed storage protein fractions. Double FISH experiment has not shown any variation in the loci number, but the positions along the chromosomes were different; both Vicia johannis and Vicia dalmatica exhibited the same interstitial 45S rRNA gene loci, while Vicia angustifolia and Vicia cordata have shown single large stretched 45S rRNA loci almost at the terminal region of the shortest chromosome. It could be concluded from the similarity matrix among the Vicia species as computed according to Jaccard coefficient from the SDS-PAGE, that V. cordata is similar to V. angustifolia and V. dalmatica by a percentage of 73 and 69%, respectively, and the most related species to V. johannis is V. dalmatica (~ 64%). ConclusionFISH and SDS-PAGE of the total seed storage proteins together reflected the similar genetic relationship among the studied species as fellows, V. angustifolia is more related to V. cordata then comes V. dalmatica and then V. johannis which is at a distal position from the other species.

Evaluation of the Putative Efficacy of a Methanolic Extract of Ocimum Basilicum in Preventing Disruption of Structural Proteins in an in Vitro System of Selenite-induced Cataractogenesis

ABSTRACTPurpose: To evaluate whether a methanolic extract of Ocimum basilicum (OB) leaves prevented lenticular protein alterations in an in-vitro model of selenite-induced cataractogenesis.Materials and Methods: Transparent lenses extirpated from Wistar rats were divided into three groups: control; selenite only; treated. Control lenses were cultured in Dulbecco’s modified Eagle’s medium (DMEM) alone, selenite only lenses were cultured in DMEM containing sodium selenite only (100 µM selenite/ml DMEM) and treated lenses were cultured in DMEM containing sodium selenite and the methanolic extract of OB leaves (200 µg of extract/ml DMEM); all lenses were cultured for 24 h and then processed. The parameters assessed in lenticular homogenates were lenticular protein sulfhydryl and carbonyl content, calcium level, insoluble to soluble protein ratio, sodium dodecyl sulphate-polyacrylamide gel electrophoretic (SDS-PAGE) patterns of lenticular proteins, and mRNA transcript and protein levels of αA-crystallin and βB1-crystallins.Results: Selenite only lenses exhibited alterations in all parameters assessed. Treated lenses exhibited values for these parameters that were comparable to those noted in normal control lenses.Conclusions: The methanolic extract of OB leaves prevented alterations in lenticular protein sulfhydryl and carbonyl content, calcium level, insoluble to soluble protein ratio, SDS-PAGE patterns of lenticular proteins, and expression of αA-crystallin and βB1-crystallin gene and proteins in cultured selenite-challenged lenses. OB may be further evaluated as a promising agent for the prevention of cataract.

Venom Composition in a Phenotypically Variable Pit Viper ( Trimeresurus insularis) across the Lesser Sunda Archipelago.

The genus Trimeresurus comprises a group of venomous pitvipers endemic to Southeast Asia and the Pacific Islands. Of these, Trimeresurus insularis, the White-lipped Island Pitviper, is a nocturnal, arboreal species that occurs on nearly every major island of the Lesser Sunda archipelago. In the current study, venom phenotypic characteristics of T. insularis sampled from eight Lesser Sunda Islands (Flores, Lembata, Lombok, Pantar, Sumba, Sumbawa, Timor, and Wetar) were evaluated via SDS-PAGE, enzymatic activity assays, fibrinogenolytic assays, gelatin zymography, and RP-HPLC, and the Sumbawa sample was characterized by venomic analysis. For additional comparative analyses, venoms were also examined from several species in the Trimeresurus complex, including T. borneensis, T. gramineus, T. puniceus, T. purpureomaculatus, T. stejnegeri, and Protobothrops flavoviridis. Despite the geographical isolation, T. insularis venoms from all eight islands demonstrated remarkable similarities in gel electrophoretic profiles and RP-HPLC patterns, and all populations had protein bands in the mass ranges of phosphodiesterases (PDE), l-amino acid oxidases (LAAO), P-III snake venom metalloproteinases (SVMP), serine proteases, cysteine-rich secretory proteins (CRISP), phospholipases A2 (PLA2), and C-type lectins. An exception was observed in the Lombok sample, which lacked protein bands in the mass range of serine protease and CRISP. Venomic analysis of the Sumbawa venom also identified these protein families, in addition to several proteins of lesser abundance (<1%), including glutaminyl cyclase, aminopeptidase, PLA2 inhibitor, phospholipase B, cobra venom factor, 5'-nucleotidase, vascular endothelial growth factor, and hyaluronidase. All T. insularis venoms exhibited similarities in thrombin-like and PDE activities, while significant differences were observed for LAAO, SVMP, and kallikrein-like activities, though these differences were only observed for a few islands. Slight but noticeable differences were also observed with fibrinogen and gelatin digestion activities. Trimeresurus insularis venoms exhibited overall similarity to the other Trimeresurus complex species examined, with the exception of P. flavoviridis venom, which showed the greatest overall differentiation. Western blot analysis revealed that all major T. insularis venom proteins were recognized by Green Pitviper ( T. albolabris) antivenom, and reactivity was also seen with most venom proteins of the other Trimeresurus species, but incomplete antivenom-venom recognition was observed against P. flavoviridis venom proteins. These results demonstrate significant conservation in the venom composition of T. insularis across the Lesser Sunda archipelago relative to the other Trimeresurus species examined.

Effect of High Pressure on the Porcine Placenral Hydrolyzing Activity of Pepsin, Trypsin and Chymotrypsin.

This study investigated the effects of protease treatments (trypsin, chymotrypsin, and pepsin) under various pressure levels (0.1-300 MPa) for the characteristics of porcine placenta hydrolysates. According to gel electrophoretic patterns, the trypsin showed the best placental hydrolyzing activity followed by chymotrypsin, regardless of the pressure levels. In particular, the peptide bands of tryptic-digested hydrolysate were not shown regardless of applied pressure levels. The peptide bands of hydrolysate treated chymotrypsin showed gradual decreases in molecular weights (Mw) with increasing pressure levels. However, the pepsin did not show any evidences of placental hydrolysis even though the pressure levels were increased to 300 MPa. The gel permeation chromatography (GPC) profiles showed that the trypsin and pepsin had better placental hydrolyzing activities under high pressure (particularly at 200 MPa), with lower Mw distributions of the hydrolysates. Pepsin also tend to lower the Mw of peptides, while the major bands of hydrolysates being treated at 300 MPa were observed at more than 7,000 Da. There were some differences in amino acid compositions of the hydrolysates, nevertheless, the peptides were mainly composed of glycine (Gly), alanine (Ala), hydroxyproline (Hyp) and proline (Pro). Consequently, the results indicate that high pressure could enhance the placental hydrolyzing activities of the selected proteases and the optimum pressure levels at which the maximum protease activity is around 200 MPa.

Dichotomous effect of caffeine, curcumin, and naringenin on genomic DNA of normal and diabetic subjects.

Nutraceutical compounds show antioxidant and prooxidant properties under stress conditions like cancer, diabetes, and other diseases. The objective of this study is to find the dichotomic behavior of caffeine, curcumin, and naringenin on DNA of diabetic and normal subjects in the presence and absence of copper, hydrogen peroxide, and complex of copper-hydrogen peroxide. Hydrogen peroxide releases hydroxyl free radicals (•OH) on oxidation of Cu (I) to Cu (II) through Fenton-type reaction to cause DNA damage. In the results, agarose gel electrophoretic pattern speculates the prooxidant effect of caffeine and antioxidant effect of curcumin on DNA in the presence of copper and hydrogen peroxide. UV-Vis spectral analysis shows hyperchromism on addition of DNA to caffeine, hypochromism with curcumin, and subtle changes with naringenin. The chosen nutraceuticals act as inducers and quenchers of oxidative free radicals arising from diabetes.

Chiral heterobimetallic complexes targeting human DNA-topoisomerase Iα

The chiral monometallic Cu(II) (1) and Zn(II) (2) and heterobimetallic Cu(II)-Sn(IV) and Zn(II)-Sn(IV) complexes with tridentate chiral Schiff base -ONO-ligand in the presence of nitrogen donor heterocyclic ligand imidazole; were prepared and characterized by various physico-chemical and spectroscopic methods. Preliminary complex-DNA interaction studies employing optical methods revealed that 3 displayed a higher propensity towards the drug target DNA double helix and recommended predominantly an electrostatic mode of interaction as well as a groove binding affinity of the complex with CT-DNA. This was quantified by Kb and KSV values of complexes 1-4, which demonstrated a multifold increase in complex 3 binding to CT DNA and clearly demonstrates its potency to act as a chemotherapeutic agent. Furthermore, the gel electrophoretic patterns of supercoiled pBR322 DNA with varying concentrations of complex 3 exhibits the ability to cleave DNA and follow a freely diffusible radical mechanism. The antiproliferative effects of complex 3 on human hepatoma cancer cells (Huh7) was investigated. Human Topo I inhibition assay by complex 3 was performed and results confirmed significantly good activity at lower concentrations than some of the classical Topo I inhibitors. Additionally, complex 3 was investigated for the expression of MMP-2 and TGF-β by real time PCR. The cellular uptake of complex 3 by HeLa cells was studied by confocal microscopy.

Optimization of enzyme-assisted extraction and characterization of collagen from Chinese sturgeon (Acipenser sturio Linnaeus) skin

Background:Sturgeon (Acipenser sturio Linnaeus) skin contains high amount of nutrients including unsaturated fatty acids and collagen. A pepsin-assisted extraction procedure was developed and optimized for the extraction of collagen from Chinese sturgeon (Acipenser sturio Linnaeus) skins.Objective:To determine the optimum conditions with the maximum yield of the pepsin-soluble collagen (PSC) extraction.Materials and Methods:The conditions of the extraction were optimized using response surface methodology. The Box–Behnken design was used to evaluate the effects of the three independent variables (extraction time, enzyme concentration, and solid–liquid ratio) on the PSC yield of the sturgeon skin.Results:The optimal conditions were: solid–liquid ratio of 1:11.88, enzyme concentration of 2.42%, and extraction time of 6.45 h. The maximum yield of 86.69% of PSC was obtained under the optimal conditions. This value was not significantly different from the predicted value (87.4%) of the RSM (P < 0.05).Conclusion:The results of this study indicated that the production of PSC from sturgeon skin is feasible and beneficial. The patterns of sodium dodecyl sulfate-polyacrylamide gel electrophoretic patterns (SDS-PAGE) indicated that the sturgeon skin contains type I collagen, which is made of α-chain and β-chain. The infrared spectra of the collagens also indicated that pepsin hydrolysis does not affect the secondary structure of collagen, especially triple-helical structure.

Zein-based polymers formed by modifications with isocyanates

Zein is a prolamine protein found in corn and has good potential in the development of industrial bioplastics. In this work, zein was modified with several isocyanates and diisocyanates in solution to evaluate its potential usage in bioplastics. The major reaction pathways were identified with the help of NMR and FTIR. For all the isocyanates employed, the surface free energy of the modified zein decreased, which phenomenon indicates decreased hydrophilicity. The moisture uptake decreased with isocyanate and diisocyanate modifications. Gel electrophoretic patterns showed that diisocyanate cross-linked a portion of zein to generate higher molecular weight species. DSC data showed single glass transition temperatures in all cases indicative that homogeneous blends were formed. The mechanical properties of modified zein were either similar to unmodified zein or reduced in some cases. Based on results from NMR and FTIR findings a reaction scheme is proposed. Our findings suggest that the reaction of zein with diisocyanates has potential usage in bioplastics formulations.

Chiral nano heterobimetallic DNA receptors: In vitro binding studies, cleavage activity and DNA condensation studies (TEM and AFM imaging)

A new chiral nano heterobimetallic Co–Sn and Zn–Sn complexes were designed and thoroughly characterized by various spectroscopic viz 1H, 13C, 119Sn NMR, ESI-MS, magnetic moment, analytical and visualization techniques. The electronic absorption and spectroscopic data reveal that the Co(II) ion exhibits a square pyramidal geometry. 119Sn NMR spectral data and powered XRD measurements support the hexa-coordinated geometry of the Sn(IV) atom. The binding studies of the heterobimetallic Co–Sn complex with CT-DNA were carried out by using various biophysical techniques viz UV–vis absorption, emission spectroscopy and cyclic voltammetric measurements, recommended predominantly electrostatic mode of interaction of the complex with CT-DNA. The gel electrophoretic patterns of supercoiled pBR322 DNA with varying concentration of the Co–Sn complex exhibit an ability to cleave DNA and follow hydrolytic pathway mechanism. Furthermore, the application of chiral nano heterobimetallic Co–Sn complex as DNA gene transporter was evaluated by DNA condensation and ascertained by employing the visualization techniques i.e. TEM and AFM, which illustrated that the Co–Sn complex induces the condensation of CT-DNA to a particulate nanostructure.

Expression of Slow Tropomyosin (TM-) in Fast Fibers of Extraocular Muscles

We previously reported marked differences in the myosin heavy and light chain (MHC and MLC) isoform composition of fast and slow fibers between the global and orbital layers of domestic dog extraocular muscles (EOM) (Bicer & Reiser, Invest. Ophthalmol. Vis. Sci. 45:138-143, 2004 and 50:157-167, 2009). In addition, many dog extraocular fibers, especially from the muscle orbital layer, have MHC and MLC isoform patterns that are distinct from those in limb skeletal muscles. The results of these studies also suggested possible differences in the tropomyosin (TM) isoform composition of fast fibers between the two EOM layers, based upon gel electrophoretic mobility patterns. The objective of this study was to determine whether differences in TM isoform expression exist between fast global and fast orbital fibers in dog EOMs. TM isoforms in global and orbital single fibers were identified by SDS-PAGE and immunoblotting. Fast and slow fibers in the global layer express fast TM-α and slow TM-γ, respectively, plus TM-β, identical to fast and slow fibers in limb muscles. Slow orbital fibers express TM-β and TM-γ, as expected. Fast orbital fibers, on the other hand, have an unusual pattern of TM-γ, along with TM-β and TM-α. The same TM isoform expression patterns were found in Sprague Dawley rat EOM fibers. Co-expression of all three TM isoforms was not observed in single fibers in limb muscles of either species. These results contribute to the understanding of the elaborate diversity in contractile protein isoform expression among mammalian EOM fibers. Supported by the National Science Foundation.

Integrity of Proteins in Human Saliva after Sterilization by Gamma Irradiation

Microbial contamination of whole human saliva is unwanted for certain in vitro applications, e.g., when utilizing it as a growth substratum for biofilm experiments. The aim of this investigation was to test gamma irradiation for its suitability to sterilize saliva and to investigate the treatment's influence on the composition and integrity of salivary proteins in comparison to filter sterilization. For inhibition of bacterial growth by gamma irradiation, a sterility assurance level of 10(-6) was determined to be reached at a dose of 3.5 kGy. At this dose, the integrity of proteins, as measured by fluorescence, circular dichroism, and gel electrophoretic banding pattern, and the enzymatic activities of salivary amylase and lysozyme were virtually unchanged. Filtration reduced the total protein concentration to about half of its original value and decreased lysozyme activity to about 10%. It can be concluded that irradiation is suitable for sterilizing whole saliva in its native form.

NUTRITIONAL CHARACTERISTICS OF LINSEED/FLAXSEED (LINUM USITATISSIMUM) AND ITS APPLICATION IN MUFFIN MAKING

ABSTRACT Flaxseed, one of the underutilized oilseeds, is a rich source of dietary fiber,ω‐3 fatty acid and protein. Flaxseed powder contained 21.8% protein and 42.7% total dietary fiber, which upon roasting reduced to 20.3 and 40.50%, respectively. Addition of flaxseed powder weakened the dough. Peak viscosity and cold paste viscosity also decreased. The volume of the muffins gradually decreased from 150 to 145 and 120 cc. Muffins prepared from 20% of raw and roasted flaxseed powder had 10.2 and 9.9% of protein and 12.8 and 11.7% of dietary fiber, respectively.ω‐3 fatty acid contents in raw and roasted flaxseed powder were 55.41 and 50.76%, and the muffins prepared with 20% raw and roasted flaxseed powder had 12.85 and 11.14%, respectively. Sodium dodecyl sulfate‐polyacrylamide gel electrophoretic pattern and gel filtration studies showed that high‐ and low‐molecular weight protein bands were identified in raw and roasted flaxseed powder and in the product.PRACTICAL APPLICATIONSFlaxseed is rich in ω‐3 fatty acid and also in fiber and protein. Also, bakery products are ready‐to‐eat products available to all people. Among the bakery products, muffins usually attract children. Hence, incorporation of flaxseed in muffins would enable to enrich the muffin with nutrients and reach the required segment of people.

Changes in SDS-polyacrylamide gel electrophoretic patterns of egg proteins during the embryonic and diapause development of the Japanese oak silkworm, Antheraea yamamai

Changes in SDS-polyacrylamide gel electrophoretic patterns of egg proteins during the embryonic and diapause development of the Japanese oak silkworm, Antheraea yamamai

Isolation and characterization of collagen from the cartilages of brownbanded bamboo shark ( Chiloscyllium punctatum) and blacktip shark ( Carcharhinus limbatus)

Acid soluble collagen (ASC) and pepsin soluble collagen (PSC) from the cartilages of brownbanded bamboo shark (BBS; Chiloscyllium punctatum) and blacktip shark (BTS; Carcharhinus limbatus) were isolated and characterized. ASC and PSC extracted from BBS cartilage showed the yields of 1.27 and 9.59 g/100 g (Based on dry weight), respectively, while yields of ASC and PSC from BTS cartilage were 1.04 and 10.30 g/100 g (based on dry weight), respectively. All collagens had protein as a major constituent with the trace amount of ash and fat. They contained glycine as the major amino acid with high contents of alanine, proline and hydroxyproline. Based on sodium dodecyl sulfate-polyacrylamide gel electrophoretic patterns and subunit compositions, all collagens more likely comprised 2 types of collagen, type I and II, and contained α- and β-chains as the major components. Peptide maps of those collagens from both species digested by V8-protease and lysyl endopeptidase were different and were completely different from those of type I collagen from calf skin. Thermal transition temperature of ASC from those collagens (36.28–36.73 °C) was slightly higher than their corresponding PSC (34.56–35.98 °C). From zeta potential analysis, isoelectric points (pI) of collagen from the cartilages of BBS and BTS were estimated to be from 6.53 to 7.03 and from 6.96 to 7.26, respectively. Fourier transform infrared (FTIR) spectra of both ASC and PSC were quite similar, suggesting that pepsin hydrolysis did not affect the secondary structure of collagen, especially triple-helical structure.

Molecular characterization of Streptococcus pyogenes from invasive disease and streptococcal toxic shock syndrome episodes

Streptococcus pyogenes causes a variety of common human diseases, including pharyngitis, scarlet fever and impetigo. Nevertheless, the past decades have witnessed a worldwide resurgence in invasive disease and streptococcal toxic shock syndrome (STSS). The objective of the present study is to evaluate the genetic diversity, virulence gene distribution (spe, sme and ssa genes) and susceptibility pattern of 10 S. pyogenes isolates causing invasive disease and STSS. The isolates were recovered from blood cultures of hospitalized patients at Hospital Santamarina and Nueva Clínica Chacabuco, Tandil, Buenos Aires, Argentina between 12/2000-04/2005. Two pulse field gel electrophoretic patterns predominated. The most frequent one included 5 characteristic isolates of emm1-T1 type, toxin gene profile speA, speB, speF, speG and smeZ. The second pattern included 2 characteristic isolates of emm3-TNT type (speB, speF, speG). The other 3 isolates corresponded to types emm49-TNT (speB, speC, speF, speG), emm75-T25 (speB, speF, speG) and emm83-TNT (speB, speF, speG, ssa, smeZ). All isolates were susceptible to penicillin, cefotaxime, erythromycin, clindamycin, chloramphenicol, tetracycline and rifampicin. The data from the present study demonstrated genetic diversity among the strains. Types emm1 and emm3 were prevalent in invasive disease. The empirical treatment with the combination of penicillin and clindamicin is still valid.

Variable starch gel electrophoretic pattern of the enzyme 6-phosphogluconate dehydrogenase in a family of donkeys (Equus asinus L.).

HereditasVolume 67, Issue 1 p. 145-146 Open Access Variable starch gel electrophoretic pattern of the enzyme 6-phosphogluconate dehydrogenase in a family of donkeys (Equus asinus L.) Håkan Bergman, Håkan BergmanSearch for more papers by this authorIngemar Gustavsson, Corresponding Author Ingemar GustavssonIngemar Gustavsson Department of Animal Genetics, Nutrition and Hygiene Royal Veterinary College S-104 05 Stockholm 50, SwedenSearch for more papers by this author Håkan Bergman, Håkan BergmanSearch for more papers by this authorIngemar Gustavsson, Corresponding Author Ingemar GustavssonIngemar Gustavsson Department of Animal Genetics, Nutrition and Hygiene Royal Veterinary College S-104 05 Stockholm 50, SwedenSearch for more papers by this author First published: March 1971 https://doi.org/10.1111/j.1601-5223.1971.tb02367.xCitations: 1AboutPDF ToolsExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat No abstract is available for this article.Citing Literature Volume67, Issue1March 1971Pages 145-146 ReferencesRelatedInformation

Linker histone‐like proteins in Muscovy duck (Cairina moschata L) erythrocyte chromatin

Linker Histone-Like proteins (LHL1 and LHL2) were identified within a linker histone complement of Muscovy duck erythrocyte chromatin. Polyacrylamide gel electrophoretic patterns of N-bromosuccinimide-cleaved LHL products as well as liquid chromatography-electrospray-ion trap mass spectrometry analyses of trypsin-digested LHL peptides revealed structural similarity of LHL1 to histone H5 and between LHL2 and histone H1 subtypes. Since the LHL proteins were stable in the presence of 2-mercaptoethanol and dithiothreitol that reduce disulfide bonds, it appeared unlikely that this doublet was a thiol-derived product of linker histones. A loss of LHL1, with a concomitant maintenance of LHL2 after treatment with dilute alkali, seems to suggest that they might represent disparate protein conjugates resulting from linker histone modifications through ester linkages.

Paraoxonase activity against nerve gases measured by capillary electrophoresis and characterization of human serum paraoxonase (PON1) polymorphism in the coding region (Q192R)

An analytical method for determining paraoxonase activity against sarin, soman and VX was established. We used capillary electrophoresis to measure directly the hydrolysis products: alkyl methylphosphonates. After enzymatic reaction of human serum paraoxonase (PON1) with nerve gas, substrate was removed with dichloromethane, and alkyl methylphoshphonates were quantified by capillary electrophoresis of reversed osmotic flow using cationic detergent and sorbic acid. This method was applied to the characterization of human serum PON1 polymorphism for nerve gas hydrolytic activity in the coding region (Q192R). PON1-192 and PON1-55 genotypes were determined by their gel electrophoretic fragmentation pattern with restriction enzymes after polymerase chain reaction (PCR) of blood leukocyte genomic DNA. Frequencies of genotypes among 63 members of our institutes with PON1-192 and PON1-55 were 9.5% ( 192QQ), 30.1% ( 192QR) and 44.4% ( 192RR), and 82.5% ( 55LL), 17.5% ( 55LM) and 0% ( 55MM), respectively. 192Q and 192R enzymes were purified from the respective genotype human plasma, using blue agarose affinity chromatography and diethyl amino ethane (DEAE) anion exchange chromatography. V max and K m were measured using Lineweaver–Burk plots for hydrolytic activities against sarin, soman and VX at pH 7.4 and 25 °C. For sarin and soman, the V max for 192Q PON1 were 3.5- and 1.5-fold higher than those for 192R PON1; and k cat/K m for 192Q PON1 were 1.3- and 2.8-fold higher than those for 192R PON1. For VX, there was little difference in V max and k cat/K m between 192Q and 192R PON1, and VX hydrolyzing activity was significantly lower than those for sarin and soman. PON1 hydrolyzed sarin and soman more effectively than paraoxon.

Effect of protein degradation on spot Mr distribution in 2‐D gels – a case study of proteolysis during development of Streptomyces coelicolor cultures

Proteolysis, a regulated biological process, is reflected by protein spot molecular weight distribution in 2-D gel electrophoretograms. Here we report studies of Streptomyces cultures as they undergo two different developmental processes involving proteolysis. Systematic changes in protein molecular weight distribution between the control samples and those with high activity of proteases were demonstrated. The observations were supported by a numerical model of degradation and its influence on the M(r) distribution. Simple statistics could be used to distinguish between normal and degradative 2-D gel electrophoretic patterns.

Keratin transamidation

Low molecular weight keratin was self-crosslinked by microbial transglutaminase (mTGase) for application to wool fabric. From amino acid determination, keratin produced by the alkaline hydrolysis of wool showed requisite glutamine and lysine required for mTGase-mediated transamidation. Keratin showed less lysyl amine content after combination with mTGase as proof of self-crosslinking. Gel electrophoretic patterns provided evidence of self-crosslinking with the development of relatively higher molecular weight protein bands within 30 min after mTGase was combined with keratin at 30 °C. Increase in the deconvoluted amide II band from IR spectra of keratin after combination with mTGase provided further evidence of transamidation. By examining the ability of keratin to self-crosslink, past findings were elucidated whereby mTGase-mediated crosslinking imparted strength to wool and keratin controlled its dimensional stability. mTGase-catalyzed isopeptide bond formation of keratin to form monosubstituted γ-amides of peptide-bound glutamine in ɛ-amino-(γ-glutamyl)lysine crosslinks. This system was effective for binding wool to wool, keratin to wool, and keratin to keratin in self-crosslinking.