UNIPROT:Q07644 - FACTA Search

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Query: UNIPROT:Q07644 (polypeptide)
72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The native and oxidized alpha-amylase inhibitor Z-2685, isolated from the culture medium of Streptomyces parvullus FH-1641, and its overlapping cleavage products were degraded by the automatic Edman technique. Digestion was carried out with pepsin, thermolysin and trypsin. The alpha-amylase inhibitor is a polypeptide consisting of 76 amino acids with a molecular mass of 8 129 Da. With the exception of methionine and lysine, all naturally occurring amino acids are present. It is interesting that identical regions exist, in particular the sequence Trp-Arg-Tyr common to all four known microbial inhibitor sequences. We believe that the side chains of these three amino acids are important for interacting with the alpha-amylase molecule. Computer alignment enabled us to show a possible binding region in the alpha-amylase molecule which might react with the inhibitors. Furthermore, homology exists to mammalian alpha-amylases. This result is explained by the assumption that the inhibitor evolved from a duplication of the original gene of the enzyme.
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PMID:The primary structure of alpha-amylase inhibitor Z-2685 from Streptomyces parvullus FH-1641. Sequence homology between inhibitor and alpha-amylase. 387 77

D-beta-Hydroxybutyrate dehydrogenase is a lipid-requiring enzyme which is localized on the inner face of the mitochondrial inner membrane. The apodehydrogenase, i.e. the purified enzyme devoid of lipid, has been purified from beef heart mitochondria and as such is inactive. It can be reactivated by insertion into phospholipid vesicles containing lecithin. Proteolytic digestion with different proteases has been carried out to obtain insight into the orientation of the enzyme in the membrane and to assess the extent of immersion of the protein into the phospholipid bilayer. Digestion of the apodehydrogenase with either trypsin, chymotrypsin, Staphylococcus aureus protease, thermolysin, carboxypeptidases A and Y, or Pronase (from Streptomyces griseus) leads to loss of activity, as assayed with phospholipid. Limited digestion with carboxypeptidase results in complete inactivation. Of the proteases tested, only Pronase and chymotrypsin cleave and inactivate the enzyme inserted into phospholipid vesicles (enzyme-phospholipid complex). For the enzyme-phospholipid complex, the loss of activity with Pronase digestion follows a single exponential decay to less than 10% of the initial activity. With chymotrypsin digestion, the staining intensity of the original approximately 31,500-dalton polypeptide decreases more rapidly than the loss of enzymic activity. The enzyme-phospholipid complex, after limited cleavage with chymotrypsin, retains enzymic activity and resonance energy transfer from protein to bound NADH and an approximately 26,000-dalton polypeptide is observed. Phospholipid alters the cleavage pattern with both chymotrypsin and Pronase, and the rate of inactivation of the enzyme-phospholipid complex is slowed in the presence of NAD(H). Moreover, the rate of inactivation of the apodehydrogenase with chymotrypsin is diminished approximately 3-fold in the presence of NAD+. Digestion of submitochondrial vesicles with either trypsin, chymotrypsin, or Pronase rapidly inactivates D-beta-hydroxybutyrate dehydrogenase; the addition of NAD+ or NADH, together with dithiothreitol and increased salt (to 50 mM), decreases the rate of inactivation, and with trypsin, virtually eliminates inactivation.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Phospholipid protection against proteolysis of D-beta-hydroxybutyrate dehydrogenase, a lecithin-requiring enzyme. 388 38

Light-mediated conformational changes in highly purified 124-kDa phytochrome preparations from etiolated oat seedlings have been identified by steric exclusion high performance liquid chromatography and limited proteolytic studies. Steric exclusion high performance liquid chromatography studies of oat and rye phytochromes show photoreversible changes in retention times, with the red absorbing form of phytochrome (Pr form) eluting later than the far red absorbing form of phytochrome produced by saturating red light illumination of Pr (Pfr form) in a variety of different mobile phase buffers. Molecular mass calibration with globular protein standards in Tris-glycol buffers provides estimates of 318-349 and 363-366 kDa for the molecular sizes of the Pr and Pfr forms, respectively. These analyses support earlier studies that phytochrome is a nonglobular homodimer of 124-kDa subunits in vitro. Limited proteolytic dissection of phytochrome in nondenaturing buffers with seven different endoproteases provides evidence for two "operational" domains within the 124-kDa subunit with molecular mass values of 69-72 and 52-55 kDa. The larger 69-72-kDa domain contains the site for the chromophore attachment as shown by gel electrophoresis derived enzyme-linked immunosorbent assay utilizing site-directed rabbit antiserum to a synthetic undecapeptide which is homologous with the chromophore binding site on oat phytochrome. This chromophore domain exhibits a compact structure, resistant to further proteolysis except near its N terminus. By contrast, the 52-55-kDa nonchromophore domain contains multiple sites for further proteolytic cleavage as revealed by rapid cleavage to smaller polypeptide fragments. Detailed kinetic analyses of the limited proteolytic cleavage of phytochrome with four endoproteases, subtilisin BPN', thermolysin, trypsin, and clostripain, has mapped specific regions within the 124-kDa subunit that participate in light-induced conformational changes. These include a 4-10-kDa region near the N terminus of the chromophore binding domain and at least two regions within the nonchromophore domain. A comprehensive peptide map of the oat phytochrome subunit is presented, which incorporates the results of these proteolytic studies with the recent, yet unpublished sequence analyses of Avena phytochrome cDNA clones which show the N-terminal localization of the chromophore binding site (Hershey, H. P., Colbert, J. T., Lissemore, J. L., Barker, R. F., and Quail, P. H. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 2332-2336).
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PMID:Structure function studies on phytochrome. Identification of light-induced conformational changes in 124-kDa Avena phytochrome in vitro. 388 93

Incubation of the neutral metalloendopeptidase thermolysin at pH 9-10 in the presence of 10 mM CaCl2 for 2 days at room temperature with subtilisin at a 50:1 molar ratio leads to a derivative possessing lower (approximately 3%) but intrinsic catalytic activity. This derivative, called thermolysin S, was isolated by gel filtration in approximately 80% yield and then separated from some residual intact thermolysin by an affinity chromatographic step on Sepharose-Gly-D-Phe. It was found that thermolysin S results from a tight association of two polypeptide fragments of apparent Mr of 24000 and 10000. Dissociation of the complex was achieved under strong denaturing conditions, such as gel filtration on a column equilibrated and eluted with 5 M guanidine hydrochloride. The positions of the clip sites were defined by amino acid analysis, end-group determination, and amino acid sequencing of the isolated fragments and shown to lie between Thr-4 and Ser-5, between Thr-224 and Gln-225, and also between Gln-225 and Asp-226. Thermolysin S, which is therefore a stable complex of fragments 5-224(225) and 225(226)-316, shows a shift in optimum pH of about 1 unit toward the acid range with respect to intact thermolysin and a Km essentially unchanged, with furylacryloyl-Gly-Leu-NH2 as substrate. Inhibitors of thermolysin such as ethoxyformic anhydride and Zn2+ ions inactivate also the nicked enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Limited proteolysis of thermolysin by subtilisin: isolation and characterization of a partially active enzyme derivative. 389 Sep 41

The amino acid sequence of an amyloid-fibril protein Es492 of immunoglobulin-lambda-light-chain origin (AL) was elucidated. The amyloid fibrils were obtained from the spleen of a patient who died from systemic amyloidosis. The amino acid sequence was elucidated from structural studies of peptides derived from digestion of the protein with trypsin, thermolysin, chymotrypsin and Staphylococcus aureus V8 proteinase and from cleavage of the protein with CNBr and BNPS-skatole. A heterogeneity in the length of the polypeptide was seen in the C-terminal region. The protein was by sequence homology to other lambda-chains shown to be of the V lambda II subgroup. Although an extensive homology was seen, some amino acid residues in positions 26, 31, 32, 40, 44, 93, 97, 98 and 99 have not previously been reported in these positions of V lambda II proteins. The significance of these residues in the fibril formation is unclear. The protein was found to contain carbohydrate, with glycosylation sites in two of the hypervariable regions.
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PMID:The amino acid sequence of a carbohydrate-containing immunoglobulin-light-chain-type amyloid-fibril protein. 393 82

An acid DNase (DNase II) from porcine spleen was purified by sequential chromatography over carboxymethyl-cellulose, blue dextran-Sepharose, hydroxylapatite, and sulfoxyethyl-cellulose. The purified enzyme shows two polypeptide bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis at Mr 35,000 (alpha chain) and 10,000 (beta chain). The sum of the two molecular weights is that of the native enzyme (45,000). Thus, the DNase II molecule is an alpha,beta dimer. The two polypeptides are not joined by disulfide bonds, but can be cross-linked chemically with dimethyl suberimidate. They are dissociable in 8 M urea, after which they can be isolated by gel filtration on Sephadex G-100, eluting with 1 M acetic acid. Once dissociated, the two polypeptides cannot be reassociated to regenerate DNase II activity. The sum of the amino acid compositions of the two polypeptides is that of the native enzyme, and both contain carbohydrate. The beta chain is devoid of histidine, half-cystine, valine, and methionine. The NH2-terminal amino acid of the alpha chain is leucine, while that of the beta chain cannot be identified by either dansylation or Edman degradation. Alkylation of an essential histidine residue of DNase II occurs on incubation of the enzyme with [2-14C] ICH2COOH (Oshima, R. G., and Price, P. A. (1973) J. Biol. Chem. 248, 7522-7526). Radioactivity is found only in the alpha chain. After hydrolysis of the alpha chain with trypsin, chymotrypsin, and thermolysin, radioactive peptides were isolated by gel filtration on Sephadex G-25 and reversed-phase high performance liquid chromatography. Sequence analyses of the radioactive peptides show alkylation of 1 of 9 histidines in the entire amino acid sequence of DNase II. The sequence around this histidine, determined by manual microsequencing and by the release of amino acids with carboxypeptidases A and B, is Ala-Thr-Glu-Asp-His-Ser-Lys-Trp.
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PMID:The subunit structure and active site sequence of porcine spleen deoxyribonuclease. 403 Jul 66

The major protein constituent of human plasma high density lipoproteins has been isolated and its complete amino-acid sequence determined. The protein, designated apolipoprotein-glutamine-I by the presence of carboxyl-terminal glutamine, is a single polypeptide chain of 245 amino-acid residues, including three residues of methionine. The protein is devoid of cysteine, cystine, and isoleucine. Cleavage of apolipoprotein-glutamine-I with cyanogen bromide yields four fragments with 94, 90, 36, and 25 amino acids. The amino-acid sequence of each fragment was determined by conventional methods, with proteolytic digestion with trypsin, chymotrypsin, and thermolysin. The alignment of the cyanogen bromide fragments was determined by the isolation of the methionine-containing tryptic peptides from apolipoprotein-glutamine-I. Inspection of the sequence of apolipoprotein-glutamine-I suggests an interesting distribution of amino acids that may account for its helical structure and its ability to bind and transport lipid.
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PMID:The primary structure of high density apolipoprotein-glutamine-I. 437 30

Bovine photoreceptor membranes have been treated with proteases to determine the accessibility of rhodopsin to these large, water soluble molecules. The polypeptides that remain associated with the membranous structure after proteolysis were detected by sodium dodecyl sulfate gel electrophoresis. Thermolysin and chymotrypsin degraded rhodopsin (apparent mol wt 35,000-36,000) to fragments of 29,000 and 23,000 apparent mol wt, respectively, without affecting the chromophoric absorption of the molecule or removing the region of the polypeptide carrying carbohydrate. The two fragments were isolated and their amino acid compositions were determined. They do not appear to be more hydrophobic than rhodopsin. Subtilisin, at low concentration and temperature, produced a fragment with the same molecular weight as that produced by thermolysin. At higher concentrations, subtilisin yields major fragments of mol wt 23,000 and 20,000 without affecting the chromophoric absorption. Two intermediate fragments of apparent mol wt 29,000 and 26,000 were detected during the course of this degradation. Carbohydrate is retained by all but the smallest fragment. Bleaching of the photoreceptor pigment did not appreciably alter any of the fragmentation patterns. Trypsin did not alter the molecular weight of rhodopsin under the conditions of this study. Approximately 35-45% of rhodopsin appears to be accessible to the aqueous environment and can be removed without affecting the chromophoric properties of the retinaldehyde-carrying region which remains bound to the membrane.
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PMID:The accessibility of bovine rhodopsin in photoreceptor membranes. 441 32

The amino acid sequence of the plastocyanin from the green alga Chlorella fusca was determined. The protein consists of a single polypeptide chain of 98 residues, and was determined by characterization of chymotryptic and thermolysin peptides. The amino acid sequence shows considerable similarity to that of higher plant plastocyanins. The protein contains a single cysteine, and the sequence in the vicinity of this residue is similar to that around the cysteine residue of bacterial azurins. The plastocyanin contains some uncharacterized carbohydrate. Detailed evidence for the sequence of the protein has been deposited as Supplementary Publication SUP 50 036 (17pp., 1 microfiche) at the British Library (Lending Division) (formerly the National Lending Library for Science and Technology), Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1973) 131, 5.
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PMID:The amino acid sequence of plastocyanin from Chlorella fusca. 446 50

The amino acid sequence of the plastocyanin from French bean (Phaseolus vulgaris) was determined. The protein consists of a single polypeptide chain of 99 residues, and the sequence was determined by characterization of CNBr, tryptic, chymotryptic and thermolysin peptides. When the sequence is compared with that from the plastocyanin of the unicellular green alga Chlorella fusca, the French-bean protein shows the deletion of the N-terminal residue, a two residue insertion and 53 identical residues. Detailed evidence for the sequence of the protein has been deposited as Supplementary Publication SUP 50037 (16pp., 1 microfiche) at the British Library (Lending Division) (formerly the National Lending Library for Science and Technology), Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1973) 131, 5.
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PMID:The amino acid sequence of plastocyanin from French bean (Phaseolus vulgaris). 446 51

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