Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.4 (trypsin)
 42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The complete amino acid sequence of D-glyceraldehyde-3-phosphate dehydrogenase from the moderate thermophile Bacillus stearothermophilus has been determined. 2. This has been achieved largely by the automated sequence analysis of large fragements derived by chemical cleavage with cyanogen bromide, BNPS-skatole [the product of reaction between N-bromosuccinimide and 2-(nitrophenyl-sulphenyl)-3-methylindole] and hydroxylamine and enzymic hydrolysis with trypsin at arginine residues. 3. The sequence is as follows: (See Text). It has been numbered to maximise homology with the four complete sequences of this enzyme from other sources. Hence the N-terminal residue is numberd 0 and two deletions and two insertions have been introduced. 4. The inability of the B. stearothermophilus apo-enzyme to transfer an acyl moiety from Cys-149 to Lys-183 oberved with muscle enzymes is explained by the replacement of lysine by arginine in the enzyme from the thermophilic organism. 5. The sequences of the S-loop regions, which form the core of the tetrameric enzyme, are similar to each other in B. stearothermophilus and Thermus aquaticus and differ from the highly conserved S-loops of three enzymes from mesophiles.
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PMID:D-glyceraldehyde-3-phosphate dehydrogenase. Complete amino-acid sequence of the enzyme from Bacillus stearothermophilus. 740 68

A 37 kDa protein that binds to diadenosine tetraphosphate (Ap4A) was purified from human HeLa cells and identified as uracil DNA glycosylase/glyceraldehyde-3-phosphate dehydrogenase (UDG/GAPDH). Utilizing photoaffinity labeling with [alpha-32P]8N3-Ap4A, an Ap4A binding protein of 37 kDa was identified from HeLa cell nuclear extracts. The 37 kDa protein was purified to homogeneity and subjected to trypsin digestion followed by amino acid sequence analysis. Two peptide sequences were determined and both had complete identity with the amino acid sequence of the 37 kDa polypeptide of UDG/GAPDH. Purified UDG/GAPDH binds to Ap4A with the same affinity as the HeLa cell nuclear 37 kDa Ap4A binding protein, and monoclonal antibodies to UDG/GAPDH cross-react with the 37 kDa Ap4A binding protein. UDG/GAPDH has been previously demonstrated to have numerous nonglycolytic activities. The UDG function is involved in DNA repair by excision of uracil from DNA. GAPDH is a RNA binding protein and binds to tRNA and AU-rich RNA. The AU-rich RNA binding has been implicated in the regulation of AU-rich element dependent mRNA turnover and translation. The identification of UDG/GAPDH as an Ap4A binding protein may be physiologically relevant to the proposed role of Ap4A as a regulatory nucleotide in cell growth.
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PMID:Uracil DNA-glycosylase/glyceraldehyde-3-phosphate dehydrogenase is an Ap4A binding protein. 762 40

Measurements of time-resolved phosphorescence anisotropy were used to monitor the rotational diffusion of eosin-labeled band 3 in membranes of the elliptocytic erythrocytes of alpacas and camels. The rotational freedom of camelid band 3 was more restricted than for human band 3. Removal of the peripheral membrane proteins from human erythrocyte membranes, by high-pH treatment, increased the band 3 rotational freedom. The same high-pH treatment of alpaca and camel erythrocyte membranes failed to alter the rotational freedom of band 3 in these species and also failed to remove ankyrin. Treatment of human and alpaca erythrocyte membranes with trypsin, which removed the cytoplasmic domain of band 3, caused a marked increase in band 3 rotational freedom in both species. We suggest that ankyrin may modulate the rotational freedom of band 3 in camelid erythrocytes, thereby influences the erythrocyte shape and deformability. The rotational freedom of band 3 in sheep, pig, and rat erythrocyte membranes was also examined and found to be slightly greater than for human band 3. This is consistent with the inability of glyceraldehyde-3-phosphate dehydrogenase to bind to band 3 in the erythrocyte membranes of these species.
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PMID:Band 3 mobility in camelid elliptocytes: implications for erythrocyte shape. 832 96

Uca pugilator serine collagenase 1 was cloned and sequenced from a fiddler crab hepatopancreas cDNA library. A full-length sequence encodes a 270-amino acid pre-pro-enzyme highly identical in structure to the chymotrypsin family of serine proteases. The zymogen form of the enzyme was expressed in Saccharomyces cerevisiae as a fusion with the alpha-factor signal sequence under control of the alcohol dehydrogenase/glyceraldehyde-3-phosphate dehydrogenase promoter. Upon activation with trypsin, the recombinant collagenase possesses collagenolytic properties identical to those of the enzyme isolated from the crab hepatopancreas. The collagenase substrate binding pocket recognizes a wide range of basic, hydrophobic, and neutral polar residues. beta-Branched and acidic amino acids are poor substrates. Acylation is rate-limiting for collagenase versus peptidyl amides, rather than deacylation, as for trypsin and chymotrypsin. Correlations relating substrate volume and hydrophobicity to catalysis were found for collagenase and compared to those for chymotrypsin and elastase. Relative enzyme efficiencies on single amino acid versus tetrapeptide amide substrates show that collagenase derives less catalytic efficiency from binding of the primary substrate residue than trypsin or chymotrypsin, but compensates in binding of the extended peptidyl residues. Serine collagenase 1 is a novel member of the chymotrypsin protease family, by virtue of its amino acid sequence and multifunctional active site.
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PMID:Substrate recognition by recombinant serine collagenase 1 from Uca pugilator. 862 18

A lambda gt11 cDNA library from Candida albicans ATCC 26555 was screened by using pooled sera from two patients with systemic candidiasis and five neutropenic patients with high levels of anti-C. albicans immunoglobulin M antibodies. Seven clones were isolated from 60,000 recombinant phages. The most reactive one contained a 0.9-kb cDNA encoding a polypeptide immunoreactive only with sera from patients with systemic candidiasis. The whole gene was isolated from a genomic library by using the cDNA as a probe. The nucleotide sequence of the coding region showed homology (78 to 79%) to the Saccharomyces cerevisiae TDH1 to TDH3 genes coding for glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and their amino acid sequences showed 76% identity; thus, this gene has been named C. albicans TDH1. A rabbit polyclonal antiserum against the purified cytosolic C. albicans GAPDH (polyclonal antibody [PAb] anti-CA-GAPDH) was used to identify the GAPDH in the beta-mercaptoethanol extracts containing cell wall moieties. Indirect immunofluorescence demonstrated the presence of GAPDH at the C. albicans cell surface, particularly on the blastoconidia. Semiquantitative flow cytometry analysis showed the sensitivity of this GAPDH form to trypsin and its resistance to be removed with 2 M NaCl or 2% sodium dodecyl sulfate. The decrease in fluorescence in the presence of soluble GAPDH indicates the specificity of the labelling. In addition, a dose-dependent GAPDH enzymatic activity was detected in intact blastoconidia and germ tube cells. This activity was reduced by pretreatment of the cells with trypsin, formaldehyde, and PAb anti-CA-GAPDH. These observations indicate that an immunogenic, enzymatically active cell wall-associated form of the glycolytic enzyme GAPDH is found at the cell surface of C. albicans cells.
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PMID:The glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase of Candida albicans is a surface antigen. 926 Sep 38

The effect of recombinant human IL-4 (rhIL-4) on the development of recombinant human stem cell factor-dependent fetal liver-derived mast cells was examined. RhIL-4 attenuates the number of mast cells that develop, preferentially affecting the MC(T) type of mast cell. Cellular levels of tryptase and chymase mRNA normalized to that of glyceraldehyde-3-phosphate dehydrogenase were not appreciably affected. Tryptase mRNA levels peaked at least 2 wk before tryptase protein and before chymase mRNA and protein, indicating that tryptase mRNA expression is an early marker of commitment to a mast cell lineage. In contrast, alpha-tryptase and beta-tryptase mRNA levels increased and decreased in parallel. The most dramatic effect of rhIL-4 was to induce expression of functional surface Fc epsilonRI. Expression was maximal by 21 days with 20 ng/ml of rhIL-4 and reached a plateau by 2 ng/ml of rhIL-4 at 4 wk. Fc epsilonRI+ cells increased modestly when myeloma IgE was added to the developing mast cells, but increased synergistically when both myeloma IgE and rhIL-4 were present together. Delayed addition of rhIL-4 progressively diminished Fc epsilonRI expression, as did withdrawal of rhIL-4 during the first 2 wk of culture. RhIL-4 selectively increased Fc epsilonRI alpha mRNA levels at least 10-fold. Mast cells developed in the presence of rhIL-4 released tryptase when exposed to anti-Fc epsilonRI alpha. In conclusion, induction of functional Fc epislonRI on recombinant human stem cell factor-dependent human fetal liver-derived mast cells by rhIL-4 harmonizes with the well-accepted ability of this cytokine to enhance IgE production by B cells.
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PMID:Effect of recombinant human IL-4 on tryptase, chymase, and Fc epsilon receptor type I expression in recombinant human stem cell factor-dependent fetal liver-derived human mast cells. 930 Jul 15

Although many abundant proteins of the postsynaptic density (PSD) are known, most of the less abundant and minor PSD proteins await identification. In this work we attempted to identify a 37 kDa protein, which represented less than 1% of the total n-octyl glucoside (NOG)-insoluble proteins, by protein sequencing. To enrich the target protein, the NOG-insoluble fraction was first electrophoresed in 6% SDS-polyacrylamide gels, and the proteins smaller than 45 kDa compressed in the gel from were electroeluted and subsequently reseparated in 10% SDS-gels. This procedure enriched the target protein to represent about 25% of the eluted proteins. Peptides were generated by digesting the target protein with trypsin directly in the gel and purified by a reverse phase high performance liquid chromatography (HPLC). Two peptides were determined for amino acid sequences. A database search revealed that both sequence were found in glyceraldehyde-3-phosphate dehydrogenase (GAPDH) with a minor discrepancy, indicating the the 37 kDa protein in the NOG-insoluble pSD fraction is an isoform of GAPDH.
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PMID:Identification of glyceraldehyde-3-phosphate dehydrogenase by protein sequencing in the rat postsynaptic density fraction. 966 75

(1) The effects of long term treatment with 3-acetylpyridine on the stability of enzymes towards heat and trypsin treatment were studied. (2) In the liver NAD or NADP provided a similar degree of protection against heat inactivation at 55 degrees C for 6-phosphogluconate dehydrogenase (24%), glyceraldehyde-3-phosphate dehydrogenase (24%) and malic enzyme (20%), low level of protection of lactate dehydrogenase (13%) but didn't affect acetylcholinesterase at all. In the muscle, however, there was substantial protection against heat inactivation by coenzyme of glyceraldehyde-3-phosphate dehydrogenase (52%), an intermediate level of protection of lactate dehydrogenase (25%), low level of protection of 6-phosphogluconate dehydrogenase (17%) and malic enzyme (17%) and almost no protection of acetylcholinesterase. (3) In the susceptibility towards trypsin a low but similar degree of protection for dehydrogenases by coenzymes was observed in the liver whereas in the muscle there was substantial protection against trypsin inactivation by NAD of glyceraldehyde-3-phosphate dehydrogenase, an intermediate level of protection of 6-phosphogluconate dehydrogenase and malic enzyme and very little protection of lactate dehydrogenase but no protection of acetylcholinesterase. Among enzymes tested, glyceraldehyde-3-phosphate dehydrogenase showed the greatest protection against heat and trypsin inactivation by NAD. (4) The results suggest that the effect of 3-acetylpyridine treatment on the stability of muscle glyceraldehyde-3-phosphate dehydrogenase appears to be quite specific and selective.
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PMID:Effects of NAD or NADP on the stability of liver and pectoral muscle enzymes in 3-acetylpyridine treated quail by heat and trypsin. 983 47

Limited proteolysis of DsbC with trypsin resulted in a compact and stable C-terminal fragment (residues 66-216), fDsbC, which retains the active site sequence, -Cys(98)-Gly-Tyr-Cys(101)-, and shows only minor differences in conformation compared with that of the intact molecule. The pK(a) of active site thiol and the K(SS) with glutathione are very close to that of DsbC, respectively; however, fDsbC is inactive as an isomerase in catalyzing the formation of correct disulfide bonds in scrambled RNase A and denatured and reduced bovine pancreatic trypsin inhibitor and shows only 13% thiol-protein oxidoreductase activity (TPOR) of DsbC. In contrast to the dimeric DsbC, fDsbC exists as a monomer and has no chaperone activity in assisting the reactivation of denatured d-glyceraldehyde-3-phosphate dehydrogenase. The heterodimer of DsbC with the inactive DsbC carboxymethylated at both active site thiols shows about 50% TPOR activity of DsbC but no isomerase activity, indicating that the DsbC subunit in the heterodimer displays full TPOR activity but little, if any, isomerase activity. It is concluded that the N-terminal sequence (residues 1-65) is essential for dimer formation and chaperone activity of DsbC. The active sites in both subunits of the dimeric DsbC appear to be essential for its isomerase activity.
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PMID:The N-terminal sequence (residues 1-65) is essential for dimerization, activities, and peptide binding of Escherichia coli DsbC. 1093 Apr 24

The stability of liver and muscle enzymes and proteins in niacin-deficient quail towards trypsin treatment in the presence and absence of coenzymes, NAD or NADP, was characterized. The protection of liver dehydrogenases by coenzymes was low when they are subjected to trypsin digestion for 60 min. In contrast, in the muscle there was substantial protection against trypsin inactivation of glyceraldehyde-3-phosphate dehydrogenase by NAD and of 6-phosphogluconate dehydrogenase by NADP. Among all enzymes tested, glyceraldehyde-3-phosphate dehydrogenase showed the greatest protection against trypsin inactivation by NAD. SDS-polyacrylamide gel electrophoresis demonstrated that muscle proteins from the niacin-deficient group were more substantially protected compared to control and pair-fed groups when liver and muscle extracts were spiked with NAD and subjected to trypsin digestion. Overall results suggest that niacin deficiency exerted specific destabilizing effects on the stability of enzymes and proteins in muscle.
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PMID:Effects of nicotinamide coenzymes on the stability of enzyme activities and proteins in niacin-deficient quail tissues against trypsin treatment. 1116 9


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