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

O-Acetylserine-O-acetylhomoserine sulfhydrylase [EC class 4.2.99], catalyzing the sulfhydrylation of both O-acetyl-L-serine (OAS) and O-acetyl-L-homoserine (OAH) (O-acetyl-L-serine(O-acetyl-L-homoserine) + H2S leads to L-cysteine (L-homocysteine) + acetate), was extracted and purified from bakers' yeast by an improved method. The purified enzyme was shown to be homogeneous on polyacrylamide gel electrophoresis both in the absence and presence of sodium dodecylsulfate and by ultracentrifugal analysis. The apo-enzyme was protected by pyridoxal phosphate (PALP) from inactivation by heat, urea, and trypsin [EC 3.4.21.4], suggesting that the binding of PALP to the apo-enzyme rendered the conformation of the protein more stable. The holo-enzyme showed absorption peaks at 420 and 330 nm due to bound PALP, in addition to a peak at 280 nm. Upon reduction with borohydride, the 420-nm peak disappeared and an increase in the 330-nm peak occurred concomitant with loss of the catalytic activity. Lysine appeared to be the pyridoxal binding site, based on identification of pyridoxyl-lysine in the hydrolyzate of the holo-enzyme. It was shown by both spectral and chemical determinations that 4 moles of PALP could bind to 200,000 g of apo-protein. The apo-enzyme showed a lower association constant with PALP than some other enzymes. Pyridoxal inhibited the activity competitively with respect to PALP. Based on these findings, it appears that the reaction mechanism of this enzyme is similar to those of other pyridoxal enzymes.
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PMID:O-Acetylserine and O-acetylhomoserine sulfhydrylase of yeast. Further purification and characterization as a pyridoxal enzyme. 79 6

The purification and characterization of two related peptides making up the glutamine binding site of formylglycinamide ribonucleotide amidotransferase from chicken liver have been presented. An amino acid residue(s) involved in binding glutamine to the enzyme was selectively labeled with [14C]iodoacetate. The labeled enzyme was reduced, carboxymethylated, and degraded by trypsin to a large radioactive peptide that yielded on acid hydrolysis only cysteine as a radioactive carboxymethylated derivative. The tryptic peptide was further digested with a protease from Streptomyces griseus. Two radioactive fractions (I and II) were obtained by gel filtration on Sephadex G-25. Furthermore, two 14C-containing peptides have been isolated from fraction I by the aid of ion exchange chromatography on AG 1-X2, AG 50W-X2 and DEAE-cellulose. Upon acid hydrolysis both peptides yielded only carboxymethylcysteine (CMCys), cystine, glycine, valine, aspartic acid, and glutamic acid. The partial sequences of the amino residues in these peptides, which are located at the glutamine binding site, have been established by the dansyl-Edman method. The sequences of amino acids of peptides a and b are Gly-Val-Cys([14C]CM)-Asp-Asx-Cys(CM)-Glx...and Gly-Val-Cys([14C]CM)-Asx-Asx..., respectively. The two peptides are undoubtedly derived from the same segment of the original protein.
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PMID:Glutamine active site of formylglycinamide ribonucleotide amidotransferase. 2. Amino acid sequence of labeled peptides. 84 8

Flagellae of Campylobacter fetus group O, types 1, 2 and 7 were prepared. First they were separated from cell bodies using an ultramix. The suspension was then centrifuged for 20 mins. at 10000 rpm and the supernatant frozen at -40 degrees C. This is a simple method for the enrichment of preparations of flagellae, as they become tangled up and accumulate in the inferior third part of the frozen liquid. The physicochemical basis of this phenomenon was discussed. After thawing and spinning for 20 mins. at 5000 rpm, the sediment was suspended in 0.9% of NaC1. The purity of the preparation was checked by electron microscopy. Antibodies to this antigen showed no cross-reaction with O-Antigen, when tested by tube agglutination. The amino acid composition of flagellae from different O-antigen serotypes was different (see Tab. 1). Cysteine could not be detected and proline only in traces. After breakdown with urea followed by gel filtration on Sephadex G 200, breakdown products of diminishing molecular size were obtained (see Fig. 2). Discelectrophoresis after ultrasonic gave 8 zones (see Fig. 3). Irrespective of serotype, thin-layer chromatography of trypsin-hydrolysed flagellae always showed 9 ninhydrin-positive spots (see Fig. 1). Only breakdown products of ultrasonic reacted with antibody. After absorption of flagellar antibody with heterologous antigen, agglutination only occurred with homologous antigen (tab. 2-5). This showed that there were different flagellar antigens. Further experiments using immunoprecipitation demonstrated two common antigenic components, a and c, and a partially common antigenic factor bb (Fig. 4), and was the basis for a classification by three groups. The three antigenic components could be separated by gel electrophoresis and detected by immunoprecipitation (Figs. 5,6).
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PMID:[Biochemical and serological investigations of flagellae of Campylobacter fetus (author's transl)]. 93 24

Nuclear basic protein from ejaculated human spermatozoa were labelled with iodo[14C1]acetic acid and fractionated by ion-exchange chromatography into several pools (named A-K). Gel electrophoresis indicated that the minor protamine components, were present in pool D and that, of the major protamine components, component 1 (pools E, F, G, H) was well separated from the unresolved mixture of component 2 and component 3 (pools I, J, K). Pools G and J were free of other contaminants. Pools D, G, and J produced different radioactive peptides on digestion with trypsin and with thermolysin, and also had quite distinct amino acid compositions. This suggests that the heterogeneity of human protamines is caused by differences in amino acid sequence. Major component 1 also seems to be heterogenous, since it was found in two distinct peaks (pools E and G), but post-translational modification as a cause of the two types of component 1 has not been ruled out. Although all the human protamine components are similar to other mammalian protamines in containing half-cysteine and tyrosine, they also have unique common features such as high histidine and high glutamic acid contents.
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PMID:The heterogeneity of the protamines from human spermatozoa. 95 98

The major form of dihydrofolate reductase from a methotrexate-resistant mutant (strain A) of Streptococcus faecium var. Durans has been purified on a large scale. Amino acid analysis of this form of the enzyme (isoenzyme 2) reveals an absence of cystine or cysteine, and sedimentation studies indicate a molecular weight of 20,800. The NH2-terminal sequence was determined by Edman degradation of the intact protein and the COOH terminus by selective tritiation and by carboxypeptidase treatment. After the action of trypsin on the citraconylated protein, seven of the expected nine peptides were purified from the digest, and after cyanogen bromide treatment of the unmodified protein, all seven of the anticipated peptides were isolated. The amino acid composition of all of these peptides has been established as well as their complete or partial sequences. From the results it was possible to order these peptides within the sequence and to establish the sequence of the NH2-terminal 60 residues and the COOH-terminal 11 residues.
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PMID:The structure of the mutant dihydrofolate reductase from Streptococcus faecium. Partial sequence and order of the limited tryptic and cyanogen bromide peptides. 115 Jun 47

The complete amino acid sequence was determined for bovine ubiquitin, and adenylate cyclase stimulating polypeptide, which is probably represented universally in living cells. Ubiquitin has a molecular weight of 8451 and consists of a single polypeptide chain containing 74 amino acid residues. It contains four arginine residues but no cysteine or trytophan residues. The first 61 amino acid residues were obtained by automated Edman degradations. Tryptic digestion of maleated ubiquitin yielded four peptide fragments that were resolved by molecular sieve chromatography and coded in order of decreasing chain length (MT-1, MT-2, MT-3, and MT-4). The automated sequenator determinations on native ubiquintin provided overlapping sequence data for three of these fragments that gave an order of MT-1, MT-3, and then MT-2; Peptide MT-4, a dipeptide, was therefore assigned to the C terminus, and the placement of peptide MT-2 was corroborated by analysis of data from carboxypeptidase digestions of maleated ubiquitin. Peptide MT-2 was domaleated and sequenced by manual Edman degradations through a single lysine residue. It was cleaved at this residue with trypsin, and the two resultant peptides were separated by ion-exchange chromatography. Manual sequencing of the C-terminal demaleated tryptic peptide of MT-2 completed the sequence of MT-2 and that of native ubiquitin. The sequence of ubiquitin was further confirmed and supported by amino acid and parital sequence anlysis of fragments obtained by digestion of maleated ubiquitin with chymotrypsin or staphylococcal protease.
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PMID:The complete amino acid sequence of ubiquitin, an adenylate cyclase stimulating polypeptide probably universal in living cells. 117 Aug 80

The amino acid sequence for vitamin D-dependent bovine intestinal calcium binding protein has been established. It contains 85 amino acids in a single chain and lacks cysteine, tryptophan, methionine, histidine, and arginine. The NH2-terminal lysine is blocked by an N-acetyl group. Enzymatic digestion with trypsin, chymotrypsin, and pepsin yielded a number of peptides which were purified by two-dimensional high voltage paper electrophoresis. These peptides were examined by end group analysis and sequenced by the dansyl procedure. The absence of tryptophan permitted by a single cleavage of the molecule by N-bromosuccinimide at the tyrosine residue at position 8 and the larger fragment was subjected to automated Edman degradation. By these means, the following sequence was established: N-Ac-Lys-Gln-Ser-Pro-Leu-Glu-Tyr-Ala-Ala-Glu-Lys-Ser-Ile-Gln-Lys-Glu-Ile-Glu-Lys-Gly-Phe-Phe-Lys-Gln-Leu-Leu-Val-Ser-Val-Gln-Lys-Ala-Gly-Asp-Lys-Glu-Ser-Leu-Gln-Pro-Leu-Phe-Thr-Leu-Leu-Lys-Ser-Gly-Pro-Glu-Glu-Asn-Leu-Lys-Glu-Ser-Gln-Asn-Gly-Pro-Asp-Leu-Ls7-Ser-Gly-Pro-Gly-Asn-Asp-Leu-Glu-Glu-Lys-Gly-Thr-Asp-Val-Phe-Ser-Leu-Lys-Gln. Microheterogeneity may exist in the molecule at residue 76 in which position threonine may be replaced by serine. Comparison of the sequence of calcium-binding protein to the "test" sequence of Tufty and Kretsinger ((1975) Science 187, 167-169) proposed to identify E-F hands in muscle proteins suggests that intestinal calcium-binding protein may likewise contain one or possibly two E-F hands which could account for calcium-binding property. Dayhoff alignment scores, however, calculated for calcium-binding protein against nine E-F hands in muscle proteins parvalbumin, troponin and alkali light chains do not indicate that intestinal calcium-binding protein is homologous to these muscle protein chains.
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PMID:Calcium-binding protein of bovine intestine. The complete amino acid sequence. 117 41

The mechanism of CO2 action in changing coccidian oocyst wall permeability was indirectly studied by substituting NO, NO2, N2O, H2S, SO2, CH4, NH3, and 8M urea in place of CO2 in an established excystation procedure. Changes in oocyst wall permeability of Eimeria stiedai, E. bovis, and E. tenella were determined by incubation in test gases and cysteine HCl followed by attempted activation of sporozoites by trypsin and bile and staining of intraoocyst components with methylene blue. The gases CH4, NO2, and N2O were negative for all 3 tests, as were SO2, NH3, and 8M urea which, in addition, were toxic to the oocysts. Both H2S and NO were capable of mimicking the action of CO2 and are related chemically to the reducing agent, and thus tend to underscore its importance in excystation. It now appears that the role of CO2 is that of an allosteric effector enhancing the action of the reducing agent.
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PMID:The effects of selected gases on excystation of coccidian oocysts. 126 26

The effect of sodium sulphite, cysteine, glutathione, mercaptoethanol and dithioerythritol (0.1-10 mmol l-1) on the activity of proteases of Microsporum gypseum was studied using azocasein, cross-linked bovine serum albumin and keratin as substrates. With the substrate without disulphide bonds (casein) no stimulation was found, and reducing agents inhibited proteolysis in most cases. With the remaining two substrates, all substances enhanced the activity of proteases probably through the cleavage of the substrate disulphide bonds. Sulphite was more effective than the four used thiols and enhanced the activity against serum albumin up to 3.2 times and against keratin up to 2.9 times. Using sulphitolysed sheep wool, keratinolytic activity increased after sulphitolysis of more than 20% of disulphide bonds. With the fully sulphitolysed wool the activity increased 43 times. The obtained results support the author's hypothesis on keratin degradation by sulphite excretion prior to attack by fungal proteases. Stimulation of proteolysis and keratinolysis by cleavage of disulphide bonds is not specific for dermatophytic proteases because trypsin and pronase behaved similarly in the experiments.
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PMID:Effect of reducing agents on proteolytic and keratinolytic activity of enzymes of Microsporum gypseum. 128 10

It has been shown that keratinase--proteinase PIV, the main enzyme of the proteolytic system of S. fradiae, is characterized by high effectiveness in its action on AcAla3OMe--exceeding elastase in catalytic effectiveness several times. This proteinase also cleaves ester and amide bonds formed by the residues of aromatic and basic amino acids, but with a lower effectiveness than chymotrypsin or trypsin. It has also been shown that proteinase IV is a typical serine enzyme highly sensitive to DFP, acting in strongly alkaline pH (about 11.2), with molecular weight 24 kDa and does not contain cysteine.
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PMID:Proteinases of Streptomyces fradiae. III. Catalytic and some physico- chemical properties of keratinolytic proteinase. 128 46


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