Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00790 (PGA)
 2,475 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pepsin was spin-labelled with N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidyl) bromoacetamide, possibly at the active site, at a beta-catboxyl group of a reactive aspartic acid. The spectrum of the spin-labelled pepsin showed that the spin probe was strongly immobilized (correlation time is greater than or equal to 10(-8) sec). Spin-labelled pepsin was thermally denatured at various temperatures and electron paramagnetic resonance (e.p.r.) spectra were taken at various times. Rates of denaturation estimated from the e.p.r. spectra at various temperatures showed that the enthalpy and entropy of thermal denaturation of spin-labelled pepsin at pH 3.5 were 48.0+/-4.9 kcal/mole and 214.7+/-14.5 e.u. respectively. Addition of conc. NaOH or 1 M acetate buffer at pH 6.0 sharpened e.p.r. spectra of the spin-labelled pepsin, indicating that the spin probe became mobilized by alkaline denaturation. Addition of urea caused unfolding of the protein which increased with the urea concentration, although only slight transition of conformational changes was observed in the e.p.r. spectra.
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PMID:Electron paramagnetic resonance studies on spin-labelling of pepsin: effects of temperature, pH and urea on its conformation. 0 80

Fermentation parameters for the production of penicillin G acylase by Escherichia coli NCIM 2400 have been evaluated. The bacterium produced the enzyme intracellularly when grown in nutrient broth containing PAA. PAA stimulated the enzyme synthesis by 8-10 fold and reduced the lag period. The optimum concentration of PAA for induction was 20 mM and addition of PAA prior to inoculation gave maximum production of PGA. Glucose, lactose, sorbitol, acetate and lactate even at 0.1% concentration catabolically repressed the enzyme formation. Peptone was the best utilised 'N' source for the enzyme production. Phosphate and yeast extract were found to be essential for both the growth and for enzyme biosynthesis. Temperature between 22-24 degrees C was optimum and under ideal condition E. coli NCIM 2400 produced 0.45-0.55 U/ml of penicillin G acylase.
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PMID:Biosynthesis of benzylpenicillin acylase by Escherichia coli NCIM-2400. 269 12

The functions of aminotelopeptide and N-terminal cross-linking of collagen I were examined. Acetic acid-soluble collagen I (ASC) was purified from neonatal bovine skin and treated with three kinds of proteases. The amino acid sequencing analysis of the N terminus showed that ASC contained a full-length aminotelopeptide. Pepsin and papain cleaved the aminotelopeptide of the alpha1 chain at the same site and the aminotelopeptide of the alpha2 chain at different sites. Proctase-treated ASC lost the whole aminotelopeptide, and the N-terminal sequence began from the tenth residue inside the triple helical region. The rates of fibril formation of pepsin-treated ASC and proctase-treated ASC were the same and were slower than that of ASC. The denaturation temperatures, monitored by CD ellipticity at 221 nm, of ASC, pepsin-treated, or papain-treated collagens were the same at 41.8 degrees C. Proctase-treated ASC showed a lower denaturation temperature of 39.9 degrees C. We also observed the morphology of the collagen fibrils under an electron microscope. The ASC fibrils were straight and thin, whereas the fibrils of pepsin-treated ASC were slightly twisted, and the fibrils from papain- and proctase-treated ASC were highly twisted and thick. When the collagen gel strength was examined by a modified method of viscosity-measurement, ASC was the strongest, followed by pepsin-treated ASC, and papain- and proctase-treated ASCs were the weakest. These results suggest that the aminotelopeptide plays important roles in fibril formation and thermal stability. In addition, the functions of intermolecular cross-linking in aminotelopeptides may contribute to the formation of fibrils in the correct staggered pattern and to strengthening the collagen gel.
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PMID:Possible involvement of aminotelopeptide in self-assembly and thermal stability of collagen I as revealed by its removal with proteases. 1085 Dec 40

A novel corrosion-resistant bioreactor composed of polyetherether ketone (PEEK), tech glass and silicium nitrite ceramics was constructed and applied for the cultivation of two newly isolated, extremely halophilic archaea producing poly(gamma-glutamic acid) (PGA), or poly(beta-hydroxy butyric acid) (PHB), respectively. These bacteria were isolated from hypersaline soil close to Aswan (Egypt). The isolate strain 40, which is related to the genus Natrialba, produced large amounts of PGA when cultivated on solid medium. Culture conditions were optimised applying the corrosion-resistant bioreactor. PGA production was dependent on NaCl concentration and occurred about at 20% (w/v) NaCl in the medium. A maximum cell density of about 1.6 g cell dry matter/l was obtained when the bioreactor was stirred and aerated in a batch fermentation process using proteose-peptone medium. The supernatant was monitored with respect to PGA formation, and after 90 h a maximum of 470 mg/l culture volume was detected by HPLC analysis. Culture conditions were optimized for the isolate 56, which accumulated PHB as intracellular granules. Batch fermentations in the stirred and aerated bioreactor applying acetate and n-butyric acid as carbon sources led to cell density of 2.28 g cell dry matter/l and a maximum PHB accumulation contributing to about 53% of cellular dry weight. About 4.6 g PHB were isolated from 10.6 g dried cells of strain 56, which exhibited a weight average molar mass of 2.3 x 10(5) g mol(-1) and a polydispersity of about 1.4.
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PMID:Polymer production by two newly isolated extremely halophilic archaea: application of a novel corrosion-resistant bioreactor. 1103 May 66

15R-Prostaglandin E2 (PGE2) methyl ester 15-acetate (1) was isolated from the R-variety of the Caribbean sea whip coral Plexaura homomalla collected in the Florida Keys. It was present in coral samples from separate collections in 2-10% of the abundance of the major prostaglandin component, PGA2 methyl ester 15-acetate. The structure of 1 was assigned based on one- and two-dimensional 1H NMR, HPLC, and LC-MS analyses. A sample of the S-variety of P. homomalla was found to contain a similar abundance of the corresponding 15S product, prostaglandin E2 methyl ester 15-acetate. The significance of PGE acetylation is discussed in relation to the proposed mechanism of PGA synthesis in the coral.
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PMID:Detection of the 15-acetate of prostaglandin E2 methyl ester as a prominent component of the prostaglandins in the gorgonian coral Plexaura homomalla. 1190 14

Selective extraction of specific cell components by enzyme or acid hydrolysis is possible from ultrathin sections for electron microscopy and parallel 2 micro sections for light microscopy of tissues fixed in formalin and embedded in a water-soluble polyepoxide, product X133/2097. Normal rat tissues fixed 15 minutes in formalin at 3 degrees C are more rapidly digested by proteinases than those fixed for the same length of time at 20 degrees C. Trypsin selectively attacks the nuclear chromatin and the ribonucleoprotein particles of the ergastroplasm, whereas mitochondria and zymogen granules resist tryptic digestion. Pepsin rapidly attacks the mitochondria and zymogen granules. The ergastoplasm and nucleus at first resist peptic digestion, but in time the entire cytoplasm and interchromatinic portion of the nucleus are attacked. Ribonuclease abolishes cytoplasmic basophilia in 2 micro sections, but parallel ultra-thin sections, stained with uranyl acetate and examined in the electron microscope, show no change in the ribonucleoprotein particles of the ergastoplasm. Desoxyribonuclease alone had no effect, but after pretreatment of the sections with pepsin or hydrochloric acid, desoxyribonuclease specifically attacked the nuclear chromatin. Nucleic acid-containing structures in the sections are gradually disintegrated by perchloric acid or hydrochloric acid.
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PMID:Ultrastructural cytochemistry. Enzyme and acid hydrolysis of nucleic acids and protein. 1376 Feb 8

The hydrolysis kinetics of native and denatured haemoglobin, using pepsin immobilized on aluminium oxide, was studied in order to produce hydrolysates containing bioactive peptides. Pepsin was immobilized on acidic alumina and on 2-ethanolamine- O -phosphate (2-EAOP)-modified acidic alumina. Surface charge of the two supports was determined as a function of pH. Kinetic studies were performed at 23 degrees C in 0.1 M acetate buffer, pH 4.5. At this pH, the surface charge of the two supports was almost the same. The coating of alumina by 2-EAOP only introduced a two carbon spacer between alumina surface and the reaction medium. Adsorption on the two supports of haemoglobin, haem and peptides produced in the course of hydrolyses were compared. Fixation of 2-EAOP on a pepsin-alumina complex gave hydrolysis kinetics of urea-denatured haemoglobin close to that obtained with the same amount of pepsin in solution, but with comparatively less adsorption of peptides and complete adsorption of haem. Heterogeneous hydrolyses of haemoglobin with pepsin, immobilized on functionalized alumina, resulted in the presence of VV-haemorphin-4, VV-haemorphin-7 and neokyotorphin in the supernatants without haem, the presence of which makes further purification difficult.
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PMID:A kinetic study of bovine haemoglobin hydrolysis by pepsin immobilized on a functionalized alumina to prepare hydrolysates containing bioactive peptides. 1503 40

Lactococcus lactis strain NZ9000(pNZpyk), which overproduces pyruvate kinase (PK), was constructed. The pNZpyk plasmid carries the P(nisA)-pyk transcriptional fusion, and the overexpression of its pyk gene was accomplished by using the nisin-inducible expression system of the NZ9000 strain. In vivo (13)C- and (31)P-NMR spectroscopy was used to evaluate the effect of this modification on the metabolism of glucose in non-growing cells. A detailed description of the kinetics of glucose, end products, glycolytic intermediates, NAD(+) and NADH was obtained. A 15-fold increase in the level of PK did not increase the overall glycolytic flux, which, on the contrary, was slightly reduced. Significant differences were observed in (i) the level of 3-phosphoglycerate (3-PGA) and phosphoenolpyruvate (PEP), metabolites associated with starvation; (ii) the rate of fructose 1,6-bisphosphate (FBP) depletion upon glucose exhaustion; and (iii) the NAD(+)/NADH ratio during glucose catabolism. In the mutant, the rate of FBP consumption after glucose depletion was notably accelerated under anaerobic conditions, whereas 3-PGA and PEP decreased to undetectable levels. Furthermore, the level of NAD(+) decreased steadily during the utilization of glucose, probably due to the unanticipated reduction in the lactate dehydrogenase activity in comparison with the control strain, NZ9000(pNZ8020). The results show that PK is an important bottleneck to carbon flux only when glucose becomes limiting; in the overproducer this constriction was no longer present, as evidenced by the faster FBP consumption and lack of accumulation of 3-PGA and PEP in anaerobic as well as aerobic conditions. Despite these clear changes, the PK-overproducing strain showed typical homolactic metabolism under anaerobic conditions, as did the strain harbouring the vector plasmid without the pyk insert. However, under an oxygen atmosphere, there was increased channelling of carbon to the production of acetate and acetoin, to the detriment of lactate production.
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PMID:Effect of pyruvate kinase overproduction on glucose metabolism of Lactococcus lactis. 1507 20

The purpose of this study was to qualitatively and quantitatively determine potential cellulose acetate butyrate (CAB) extractables in a way to meaningfully predict the in vivo exposure resulting from clinical administration. Extractions of CAB-381-20 were performed in several solvent systems, consistently resulting in the detection of three extractables. The extractables have been identified as acetic acid, butyric acid, and E-2-ethyl-2-hexenoic acid (E-EHA) by LC/UV, LC/MS and NMR. Extraction studies of CAB powders in acetonitrile/phosphate buffer demonstrated quantitative extraction in 1 h for acetic acid (approximately 150 microg/g), butyric acid (approximately 200 microg/g), and EHA (approximately 20 microg/g). Subsequently, extraction studies for CAB powders and coated tablets in USP simulated gastric and intestinal fluids were performed to evaluate potential in vivo exposure. Similarly, acetic and butyric acids were quantitatively extracted from CAB-381-20 powder after 24 h exposure in both USP simulated fluids. The amounts of EHA extracted from CAB powder after 24 h were determined to be 2 and 16 microg/g in USP simulated gastric and intestinal fluids, respectively. After 24 h exposure in USP simulated fluids, the maximum amount of EHA extracted corresponds to < 0.3 microg of EHA per tablet. Pepsin and pancreatin in USP simulated fluids had no effect on EHA extraction and quantitation.
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PMID:Identification and quantitation of extractables from cellulose acetate butyrate (CAB) and estimation of their in vivo exposure levels. 1519 22

Bacteria in a biofilm are enmeshed in a self-synthesized extracellular polysaccharide matrix that holds the bacteria together in a mass and firmly attaches the bacterial mass to the underlying surface. A major component of the extracellular polysaccharide matrix in several phylogenetically diverse bacteria is PGA, a linear polymer of N-acetylglucosamine residues in beta(1,6)-linkage. PGA is produced by the Gram-negative periodontopathogen Actinobacillus actinomycetemcomitans as well as by the Gram-positive device-associated pathogen Staphylococcus epidermidis. We recently reported that A.actinomycetemcomitans produces a soluble glycoside hydrolase named dispersin B, which degrades PGA. Here, we present the crystal structure of dispersin B at 2.0A in complex with a glycerol and an acetate ion at the active site. The enzyme crystallizes in the orthorhombic space group C222(1) with cell dimensions a=41.02A, b=86.13A, c=185.77A. The core of the enzyme consists a (beta/alpha)(8) barrel topology similar to other beta-hexosaminidases but significant differences exist in the arrangement of loops hovering in the vicinity of the active site. The location and interactions of the glycerol and acetate moieties in conjunction with the sequence analysis suggest that dispersin B cleaves beta(1,6)-linked N-acetylglucosamine polymer using a catalytic machinery similar to other family 20 hexosaminidases which cleave beta(1,4)-linked N-acetylglucosamine residues.
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PMID:Structural analysis of dispersin B, a biofilm-releasing glycoside hydrolase from the periodontopathogen Actinobacillus actinomycetemcomitans. 1587 75


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