Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.27 (thermolysin)
 1,894 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The low molecular weight, glutamine-rich storage protein isolated from the seeds of Ricinus communis (castor beans) has been shown to consist of two different polypeptide chains linked by disulfide bond(s). The small subunit is composed of 34 amino acids with a proline at its NH2 terminus, whereas the large subunit contains 61 amino acids with a cyclized glutamine as the NH2-terminal residue. The complete amino acid sequence of both subunits has been determined through characterization of the isolated subunits and selected peptides from trypsin, chymotrypsin, thermolysin, and cyanogen bromide cleavage. The intact protein possesses a large number of glutaminyl and half-cystinyl residues and exhibits sequence heterogeneity as observed from peptide sequences. Comparison of the sequence of this protein and those of other seed proteins indicates some structural similarities between them. The amino acid sequences of the two polypeptide chains of castor bean storage protein are: (formula, see text).
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PMID:Amino acid sequence of small and large subunits of seed storage protein from Ricinus communis. 717 64

1-Azido[3H]pyrene ([3H]AP) has been synthesized with high specific radioactivity (3 Ci/mmol) and used to photochemically label retinal rod outer segment disk membranes. The reagent reacts with rhodopsin and a Mr approximately 240000 protein as well as with membrane lipids. When [3H]AP-rhodopsin is digested with thermolysin in the disk membrane, both membrane-bound fragments of rhodopsin, F1 and F2, are found to contain [3H]AP. Reaction of the reagent appears to be restricted to the lipophilic surface of rhodopsin inasmuch as the presence of the nitrene scavenger glutathione in the aqueous medium does not significantly reduce 3H incorporation into rhodopsin. Labeled F1 and F2 were prepared, their cyanogen bromide peptides partially separated, and specific radioactivities determined. A factor of 4.4-fold in specific radioactivities of peptide pools was found, which suggests that some specificity has been shown in the reaction of [3H]AP toward different surfaces of rhodopsin.
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PMID:Topography of rhodopsin in retinal rod outer segment disk membranes. Photochemical labeling with 1-azidopyrene. 723 11

The complete amino acid sequence of copper-zinc superoxide dismutase from horse liver is reported. The molecule consists of 153 amino acids and has a Mr = 16,000. The primary structure was determined by automated and manual sequence analysis on fragments produced by cleavage of the S-carboxymethylated protein with cyanogen bromide and on peptides obtained by digestion with trypsin, thermolysin, Staphylococcus aureus protease, or subtilisin. The protein is devoid of tryptophan and tyrosine and displays an acetylated NH2 terminus. Comparison of its primary structure with the known sequences of copper-zinc superoxide dismutases from bovine and human erythrocytes and from yeast reveals a high degree of sequence homology among the four enzymes. This is especially borne out in the regions containing the amino acid residues involved in the metal binding and the half-cystine residues forming the intramolecular disulfide bridge. The striking conservation of the preponderant glycine residues known to be important for the pronounced protein folding in bovine erythrocyte superoxide dismutase suggests similar three-dimensional structures for human erythrocyte, horse liver, and yeast copper-zinc superoxide dismutases.
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PMID:Amino acid sequence of copper-zinc superoxide dismutase from horse liver. 729 16

The M protein of influenza is the predominant structural component of the virus. The interactions of this protein with the viral lipid or with other proteins are not known. The ability of M to interact with viral or other lipids was investigated. Purified M was mixed with viral lipid or egg phosphatidylcholine and was incorporated into vesicles (i) by addition of sodium deoxycholate followed by dialysis or (ii) by sonication. Between 90 and 100% of the M became firmly associated with the lipid by either of these two methods, whereas nucleoprotein failed to associate with the vesicles. From association also occurred if M was mixed with performed vesicles. Most of the M attached to the vesicles could be hydrolyzed with proteolytic enzymes such as trypsin or thermolysin, except for a small fragment of about 5,000 daltons which remained associated with the lipid vesicles. The ability of fragments of M to interact with lipids was also investigated. Of 13 fragments produced by cleavage with cyanogen bromide, 3 specifically associated with lipid vesicles. The data indicate that a specific portion of the M molecule has a high affinity for lipid bilayers of various origins.
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PMID:Interaction of influenza M protein with viral lipid and phosphatidylcholine vesicles. 743 87

Myoglobin isolated from red muscle of the gummy shark M. antarcticus was purified by gel filtration and ion-exchange chromatography on carboxymethyl cellulose in 8 M urea-thiol buffer. Amino acid analysis and sequence determination showed 148 amino acid residues. The amino terminal residue is acetylated as shown by nuclear magnetic resonance and mass spectrographic analysis of an N-terminal peptide. There is a deletion of four residues at the amino terminal end as well as one residue in the CD interhelical area relative to other myoglobins. These overall differences were also found previously in myoglobin of Heterodontus portusjacksoni. The complete amino acid sequence has been determined following digestion with trypsin, chymotrypsin, thermolysin, staphylococcal protease and cyanogen bromide. Sequences of purified peptides were determined by the dansyl-Edman procedure. The amino acid sequence showed approximately 88 differences from mammalian, monotreme, bird and tuna myoglobins, slightly more than previously reported for H. portusjacksoni usually considered a more primitive animal. There were 24 residues common to both shark myoglobins that were different from those present in other myoglobins. The sequence has been compared to the myoglobin of yellowfin tuna and other myoglobins.
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PMID:Myoglobins of cartilaginous fishes. II. Isolation and amino acid sequence of myoglobin of the shark Mustelus antarcticus. 743 64

The activity of thermolysin is greatly enhanced in the presence of high concentrations of neutral salts [Holmquist, B. and Vallee, B.L. (1976) Biochemistry 15, 101-107; Inouye, K. (1992) J. Biochem. 112, 335-340]. NaBr and NaCl are the most effective for the activation. An absorption difference spectrum with a peak around 293 nm, which is characteristic of the red-shift of a tryptophyl residue caused by charge effects, was observed on mixing of thermolysin with NaCl. As the peak disappeared in the presence of competitive inhibitors of the enzyme (phosphoramidon and zincov), it was considered to be derived from a tryptophyl residue (Trp 115) located in the active site of the enzyme. On the other hand, this peak was not observed on the mixing of thermolysin and NaBr, indicating that the slight difference in size between chloride and bromide ions is critical for the interaction with the tryptophyl residue. NaCl and NaBr exhibit comparable effects on the activation of thermolysin regardless of the considerable discrepancy in their effects on the absorptivity difference around 293 nm. This suggests that the interaction of salts with Trp 115 is not necessarily correlated with the activation of thermolysin.
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PMID:A spectrophotometric study on the interaction of thermolysin with chloride and bromide ions, and the state of tryptophyl residue 115. 785 70

A purification procedure for guanylate kinase from pig brain has been developed consisting of ammonium sulfate precipitation and heptane extraction of the crude extract, hydrophobic-interaction chromatography, affinity chromatography and chromatofocussing. From 1.75 kg pig brain, 1.2 mg enzyme was isolated with a yield of 18% and a purity of about 90%. For sequence determination, the protein was cleaved with trypsin, cyanogen bromide and endoproteinase Glu-C. Some of the isolated peptides were subcleaved with chymotrypsin, thermolysin or trifluoroacetic acid. The blocked N-terminus was analyzed by mass spectrometry and by amino acid analysis of a tryptic peptide, while the C-terminus was found in a tryptic and a chymotryptic peptide and confirmed by a carboxypeptidase Y digestion. The sequence contains 197 amino acids with a M(r) of 21,831, one tryptophan and one cysteine residue. It has been compared to those of the homologous enzymes of yeast and Escherichia coli, as well as to proteins from sequence data banks that show similarities. The sequence is discussed in the light of the known spatial structure of yeast guanylate kinase.
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PMID:Purification and sequence determination of guanylate kinase from pig brain. 809 61

The effect of sodium halide salts on the hydrolysis of three of the dansyl (Dns) peptide substrates described in the previous paper (Yang & Van Wart, 1994) by thermolysin have been studied. Increasing concentrations of sodium chloride decrease the KM value for the hydrolysis of the tripeptides Dns-Gly-Phe-Ala and Dns-Ala-Phe-Ala but leave kcat unaltered. This kinetic behavior is described by a nonessential activation mechanism in which chloride binds preferentially to the enzyme-substrate complex. Similar trends are found for the sodium bromide and fluoride salts. In contrast, sodium chloride decreases both KM and kcat almost equally for the hydrolysis of Dns-Ala-Ala-Phe-Ala, leaving kcat/KM unchanged. Thus, chloride is an uncompetitive inhibitor of this substrate. Molecular modeling studies have been carried out in order to explain the effect of chloride on the binding of these dansyl peptides. The decrease in KM for the hydrolysis of all three substrates is attributed to an interaction of chloride with Arg-203 located in the active site to stabilize the enzyme-substrate complexes. The differential effect of chloride on the kcat values for the hydrolysis of the dansyl tripeptides vs dansyl tetrapeptide is related to differences in binding on the Pn side of the substrates. The tripeptides are predicted to bind to the active site of thermolysin in a single low-energy conformation. However, there are two populations of low-energy binding modes for the tetrapeptide, one of which is believed to be a more productive binding mode.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Differential effect of halide anions on the hydrolysis of different dansyl substrates by thermolysin. 820 86

The complete amino acid sequence of the Fd' region including the VH part, the CH1 domain, and the hinge segment of the biologically relevant monoclonal mouse anti-alpha (2-8) polysialic acid antibody mAb735 is presented. The reduced and carboxymethylated H-chain was digested with trypsin and cyanogen bromide. For subfragmentation selected peptides were cleaved with thermolysin and endoproteinase Asp-N. The generated peptides were isolated by RP-HPLC and characterized by sequence analysis, plasma desorption mass spectrometry (PDMS), and amino acid analysis. The N-terminal sequence was determined after enzymatic deprotection with pyroglutamate aminopeptidase. According to Kabat et al. the variable region of the H-chain belongs to the subgroup II. Sequence data from the constant region indicate that mAb735 represents the gamma 2a isotype.
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PMID:Primary structure of the murine monoclonal IgG2a antibody mAb735 against alpha (2-8) polysialic acid. 2. Amino acid sequence of the heavy (H-) chain Fd' region. 829 1

In this study, we sequenced a new type I ribosome-inactivating protein, trichoanguina, from the seeds of Trichosanthes anguina (snake gourd). Trichoanguina is a basic glycoprotein having an apparent molecular mass of 35.0 kD and possessing strong ribosome-inactivating activity. Trichoanguina was cleaved with cyanogen bromide and partially digested with thermolysin, chymotrypsin, trypsin and Staphylococcus aureus V8 protease. The subsequent peptide fragments were separated by SDS-polyacrylamide gel electrophoresis, followed by electroblotting to polyvinylidene difluoride membranes and then sequencing. The sequencing of trichoanguina was completed, consisting of 245 amino acid residues. The sequencing of trichoanguina revealed a considerable homology to trichosanthin and alpha-trichosanthin, which are known as abortifacient, ribosome-inactivating and antihuman immunodeficiency virus proteins, with 46.7% and 55.6% amino acid identities, respectively. The sequence conserves two active sites: Glu-158 and Arg-161. Copyright 1996 S. Karger AG, Basel
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PMID:Amino Acid Sequence of Trichoanguina, a Ribosomal-Inactivating Protein from Trichosanthes anguinea Seeds. 1172 98


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