scholarly journals Amino acid sequence of HSP-1, a major protein of stallion seminal plasma: effect of glycosylation on its heparin- and gelatin-binding capabilities

1995 ◽  
Vol 310 (2) ◽  
pp. 615-622 ◽  
Author(s):  
J J Calvete ◽  
K Mann ◽  
W Schäfer ◽  
L Sanz ◽  
M Reinert ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Seminal Plasma ◽  
Binding Activity ◽  
Size Exclusion ◽  
Major Protein ◽  
Heparin Binding ◽  
Aggregation State ◽  
Consensus Pattern

We report the complete amino acid sequence of HSP-1, a major protein isolated from stallion seminal plasma or acid extracts of ejaculated spermatozoa. The protein consists of 121 amino acids organized in two types of homologous repeats arranged in the pattern AA‘BB’. Each of the 13-15-residue A-type repeats contains two O-linked oligosaccharide chains. The B-type repeats span 44-47 amino acids each, are not glycosylated, and have the consensus pattern of the gelatin-binding fibronectin type-II module. This domain also occurs in the major bovine seminal plasma heparin-binding proteins PDC-109 (BSP-A1/A2) and BSP-A3. However, unlike the bovine proteins which bind quantitatively to a heparin-Sepharose column, stallion HSP-1 was recovered in both the flow-through and the heparin-bound fractions. Structural analysis showed that the two HSP-1 forms contain identical polypeptide chains which are differently glycosylated. Moreover, size-exclusion chromatography showed that heparin-bound HSP-1 associates with HSP-2, another major seminal plasma protein, into a 90 kDa product, whereas the non-heparin-bound glycoform of HSP-1 is eluted as a monomeric (14 kDa) protein. This suggests that glycosylation may have an indirect effect on the heparin-binding ability of HSP-1 through modulation of its aggregation state. On the other hand, both glycoforms of HSP-1 displayed gelatin-binding activity, indicating that the molecular determinants for binding heparin and gelatin are different.

Aggregated Forms of Bull Seminal Plasma Proteins and Their Heparin-Binding Activity

2004 ◽  
Vol 69 (3) ◽  
pp. 616-630 ◽  
Author(s):  
Petra Jelínková ◽  
Helena Ryšlavá ◽  
Jiří Liberda ◽  
Věra Jonáková ◽  
Marie Tichá
Keyword(s):  
Molecular Weight ◽  
Seminal Plasma ◽  
Plasma Proteins ◽  
Sodium Dodecyl ◽  
Reversed Phase ◽  
Binding Activity ◽  
Size Exclusion ◽  
Heparin Binding ◽  
Aggregation State ◽  
Seminal Plasma Proteins

Heparin-binding activity of bull seminal plasma proteins was shown to be dependent on their aggregation state. The protein fraction interacting with immobilized heparin was characterized by large polydispersity in the region of molecular weight of 60 000-10 000, while that not retained on the affinity carrier was present as aggregates with molecular weight >100 000. Components of heparin-binding and non-heparin-binding fractions were separated by RP HPLC (reversed-phase HPLC) and analyzed by SDS (sodium dodecyl sulfate) electrophoresis and N-terminal sequencing. Size exclusion chromatography of whole seminal plasma and heparin-binding proteins in the presence of D-fructose (as a component of seminal plasma) showed that the region of molecular weights of protein-associated forms was shifted to lower values. An increase of heparin-binding activity of bull proteins, as determined by ELBA (Enzyme-Linked Binding Assay), correlates with a decrease of their aggregation state. The modulation of the aggregation state of bull proteins by seminal plasma components and, in this way, also of their heparin-binding properties suggests possible mechanisms for capacitation mediated by these proteins.

scholarly journals Cloning, sequence, expression and characterization of human beta-mannosidase.

2008 ◽  
Vol 55 (3) ◽  
pp. 479-490 ◽  
Author(s):  
Zahoor Qadir Samra ◽  
Muhammad Amin Athar
Keyword(s):  
Amino Acids ◽  
Enzyme Activity ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Dna Sequence ◽  
Dna Sequences ◽  
Blot Analysis ◽  
Size Exclusion ◽  
Expression Vectors

Beta-mannosidase (EC 3.2.1.25, MANB) dissects the non-reducing end of N-linked mannose moieties of glycoproteins in eukaryotic cells. The human beta-mannosidase gene was amplified by RT-PCR, cloned and sequenced. The DNA sequence was compared with reported human beta-mannosidase DNA sequence and sixteen nucleotide differences were found. The deduced amino-acid sequence showed that seven codons coded the same amino acids and nine codons coded different amino acids with reference to nucleotide substitution positions but did not affect recombinant MANB enzyme activity. No splice mutation was observed after comparison with reported MANB DNA sequences. A 75% homology of deduced amino-acid sequence was observed with mouse, goat and bovine beta-mannosidase amino-acid sequences. The cloned beta-mannosidase gene was subcloned into pET22b+ and pET28a+ expression vectors to transform the BL21-codon plus cells for expression of recombinant MAN22 and MAN28 enzymes, respectively. The optimized conditions for overexpression of recombinant beta-mannosidase enzyme were induction with 1 mM IPTG for 12 h at 37 degrees C. The expressed beta-mannosidase enzyme was purified to homogeneity by a combination of DEAE-ion exchange and size exclusion chromatography. The molecular mass of MAN22 and MAN28 enzymes is 97 kDa by SDS/PAGE and is confirmed by western blot analysis. The recombinant enzymes are active at 37 degrees C and at pH 5.0 and showed activity with p-nitrophenyl-beta-d-mannopyranoside and not with p-nitrophenyl-alpha-d-mannopyranoside. The K(m) value of enzymes was 2.53 mM. The enzyme activity was inhibited by Zn(2+), Co(2+), Cu(2+), Pb(2+), Ag(1+), iodoacetate, SDS, DMF, DMSO and ethanol. Fe(3+), Ca(2+) Mg(2+), Mn(2+), Triton X-100 and PMSF did not inhibit the enzyme activity. Northern blot analysis showed a transcript of about 3.7 kb in all cells and tissues studied. This is the first report on the expression and characterization of recombinant human MANB enzyme.

scholarly journals Complete amino acid sequence of BSP-A3 from bovine seminal plasma. Homology to PDC-109 and to the collagen-binding domain of fibronectin

1987 ◽  
Vol 243 (1) ◽  
pp. 195-203 ◽  
Author(s):  
N G Seidah ◽  
P Manjunath ◽  
J Rochemont ◽  
M R Sairam ◽  
M Chrétien
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Seminal Plasma ◽  
Complete Characterization ◽  
Binding Domain ◽  
Collagen Binding ◽  
Ancestral Gene ◽  
Complete Amino Acid Sequence ◽  
Molecular Variants

Bovine seminal plasma was shown to contain three similar proteins, called BSP-A1, BSP-A2 and BSP-A3. Both BSP-A1 and BSP-A2 were shown to be molecular variants of a recently characterized peptide called PDC-109. They seem to differ only in their degree of glycosylation and otherwise seem to possess an identical amino acid composition. The work in the present paper deals with the complete characterization of the third member of this series, namely BSP-A3. The complete amino acid sequence revealed that it is composed of 115 amino acids and predicts a Mr of 13,403. An analysis of the primary structure of BSP-A3 revealed a high degree of internal homology, with two homologous domains composed of 39 (residues 28-66) and 43 (residues 73-115) amino acids. An exhaustive computer-bank search for the similarity of this sequence to any known protein, or segment thereof, revealed two significant homologies. The first is between PDC-109 and BSP-A3, which is so high that we can confidently predict that both proteins evolved from a single ancestral gene. The collagen-binding domain of bovine fibronectin (type II sequence) was also found to be highly homologous to both BSP-A3 and PDC-109.

Complete Covalent Structure of Human Platelet Factor 4

1979 ◽  
Vol 42 (05) ◽  
pp. 1652-1660 ◽  
Author(s):  
Francis J Morgan ◽  
Geoffrey S Begg ◽  
Colin N Chesterman
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Human Platelet ◽  
Platelet Factor 4 ◽  
Platelet Factor ◽  
Disulphide Bonds ◽  
Covalent Structure

SummaryThe amino acid sequence of the subunit of human platelet factor 4 has been determined. Human platelet factor 4 consists of identical subunits containing 70 amino acids, each with a molecular weight of 7,756. The molecule contains no methionine, phenylalanine or tryptophan. The proposed amino acid sequence of PF4 is: Glu-Ala-Glu-Glu-Asp-Gly-Asp-Leu-Gln-Cys-Leu-Cys-Val-Lys-Thr-Thr-Ser- Gln-Val-Arg-Pro-Arg-His-Ile-Thr-Ser-Leu-Glu-Val-Ile-Lys-Ala-Gly-Pro-His-Cys-Pro-Thr-Ala-Gin- Leu-Ile-Ala-Thr-Leu-Lys-Asn-Gly-Arg-Lys-Ile-Cys-Leu-Asp-Leu-Gln-Ala-Pro-Leu-Tyr-Lys-Lys- Ile-Ile-Lys-Lys-Leu-Leu-Glu-Ser. From consideration of the homology with p-thromboglobulin, disulphide bonds between residues 10 and 36 and between residues 12 and 52 can be inferred.

SOME ASPECTS OF THE STRUCTURE OF HEMOGLOBIN

1964 ◽  
Vol 42 (6) ◽  
pp. 755-762 ◽  
Author(s):  
David B. Smith
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Partial Amino Acid Sequence ◽  
Heme Pocket ◽  
Terminal Amino ◽  
Polypeptide Chains ◽  
Kinetics Of ◽  
Β Chain

An outline of present ideas concerning the arrangement, folding, and chemistry of the polypeptide chains of hemoglobin is given with some references to present know ledge of myoglobin.New material includes a partial amino acid sequence of the β-chain of horse hemoglobin, details concerning the amino acids lining the heme pocket of horse hemoglobin, and the effects of carboxypeptidases A and B on horse oxy- and horse deoxy-hemoglobin. The kinetics of the latter reactions are not simple. The C-terminal amino acids are released more rapidly from the oxygenated form.

scholarly journals The amino acid sequence of cytochromes c-551 from three species of Pseudomonas

1973 ◽  
Vol 131 (3) ◽  
pp. 485-498 ◽  
Author(s):  
R. P. Ambler ◽  
Margaret Wynn
Keyword(s):  
Amino Acids ◽  
Pseudomonas Aeruginosa ◽  
Amino Acid ◽  
Cytochrome C ◽  
Amino Acid Sequence ◽  
Amino Acid Sequences ◽  
Mitochondrial Cytochrome ◽  
Cytochromes C ◽  
Lending Library ◽  
Single Peptide

The amino acid sequences of the cytochromes c-551 from three species of Pseudomonas have been determined. Each resembles the protein from Pseudomonas strain P6009 (now known to be Pseudomonas aeruginosa, not Pseudomonas fluorescens) in containing 82 amino acids in a single peptide chain, with a haem group covalently attached to cysteine residues 12 and 15. In all four sequences 43 residues are identical. Although by bacteriological criteria the organisms are closely related, the differences between pairs of sequences range from 22% to 39%. These values should be compared with the differences in the sequence of mitochondrial cytochrome c between mammals and amphibians (about 18%) or between mammals and insects (about 33%). Detailed evidence for the amino acid sequences of the proteins has been deposited as Supplementary Publication SUP 50015 at the National Lending Library for Science and Technology, Boston Spa, Yorks. LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1973), 131, 5.

Conserved DNA binding and self-association domains of the Drosophila zeste protein

1992 ◽  
Vol 12 (2) ◽  
pp. 598-608
Author(s):  
J D Chen ◽  
C S Chan ◽  
V Pirrotta
Keyword(s):  
Amino Acid ◽  
Dna Binding ◽  
Amino Acid Sequence ◽  
Coiled Coil ◽  
Helical Structure ◽  
Binding Activity ◽  
Binding Domain ◽  
Drosophila Virilis ◽  
Predicted Amino Acid Sequence ◽  
Dna Binding Domain

The zeste gene product is involved in two types of genetic effects dependent on chromosome pairing: transvection and the zeste-white interaction. Comparison of the predicted amino acid sequence with that of the Drosophila virilis gene shows that several blocks of amino acid sequence have been very highly conserved. One of these regions corresponds to the DNA binding domain. Site-directed mutations in this region indicate that a sequence resembling that of the homeodomain DNA recognition helix is essential for DNA binding activity. The integrity of an amphipathic helical region is also essential for binding activity and is likely to be responsible for dimerization of the DNA binding domain. Another very strongly conserved domain of zeste is the C-terminal region, predicted to form a long helical structure with two sets of heptad repeats that constitute two long hydrophobic ridges at opposite ends and on opposite faces of the helix. We show that this domain is responsible for the extensive aggregation properties of zeste that are required for its role in transvection phenomena. A model is proposed according to which the hydrophobic ridges induce the formation of open-ended coiled-coil structures holding together many hundreds of zeste molecules and possibly anchoring these complexes to other nuclear structures.

scholarly journals Amino acid sequence of the human fibronectin receptor.

1987 ◽  
Vol 105 (3) ◽  
pp. 1183-1190 ◽  
Author(s):  
W S Argraves ◽  
S Suzuki ◽  
H Arai ◽  
K Thompson ◽  
M D Pierschbacher ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Calcium Binding ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Receptor Subunit ◽  
Fibronectin Receptor ◽  
Adhesion Receptor ◽  
Receptor Beta

The amino acid sequence deduced from cDNA of the human placental fibronectin receptor is reported. The receptor is composed of two subunits: an alpha subunit of 1,008 amino acids which is processed into two polypeptides disulfide bonded to one another, and a beta subunit of 778 amino acids. Each subunit has near its COOH terminus a hydrophobic segment. This and other sequence features suggest a structure for the receptor in which the hydrophobic segments serve as transmembrane domains anchoring each subunit to the membrane and dividing each into a large ectodomain and a short cytoplasmic domain. The alpha subunit ectodomain has five sequence elements homologous to consensus Ca2+-binding sites of several calcium-binding proteins, and the beta subunit contains a fourfold repeat strikingly rich in cysteine. The alpha subunit sequence is 46% homologous to the alpha subunit of the vitronectin receptor. The beta subunit is 44% homologous to the human platelet adhesion receptor subunit IIIa and 47% homologous to a leukocyte adhesion receptor beta subunit. The high degree of homology (85%) of the beta subunit with one of the polypeptides of a chicken adhesion receptor complex referred to as integrin complex strongly suggests that the latter polypeptide is the chicken homologue of the fibronectin receptor beta subunit. These receptor subunit homologies define a superfamily of adhesion receptors. The availability of the entire protein sequence for the fibronectin receptor will facilitate studies on the functions of these receptors.

scholarly journals Crotalase, a Fibrinogen-Clotting Snake Venom Enzyme: Primary Structure and Evidence for a Fibrinogen Recognition Exosite Different from Thrombin

1999 ◽  
Vol 81 (01) ◽  
pp. 81-86 ◽  
Author(s):  
Agnes Henschen-Edman ◽  
Ida Theodor ◽  
Brian Edwards ◽  
Hubert Pirkle
Keyword(s):  
Molecular Weight ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Spatial Modeling ◽  
Glycosylation Site ◽  
Amino Sugars ◽  
Edman Degradation ◽  
Heparin Binding ◽  
Heparin Binding Site ◽  
Total Molecular Weight

SummaryCrotalase, a fibrinogen-clotting enzyme isolated from the venom of Crotalus adamanteus, and its overlapping fragments were subjected to Edman degradation. The resulting amino acid sequence, VIGGDEC NINEHRFLVALYDYWSQLFLCGGTLINNEWVLTAAHCDRTHI LIYVGVHDRSVQFDKEQRRFPKEKYFFDCSNNFTKWDKDIM LIRLNKPVSYSEHIAPLSLPSSPPIVGSVCRAMGWGQTTSPQET LPDVPHCANINLLDYEVCRTAHPQFRLPATSRTLCAGVLEG GIDTCNRDSGGPLICNGQFQGIVFWGPDPCAQPDKPGLYTK VFDHLDWIQSIIAGEKTVNCP, is characteristic of a serine protein-ase. Comparison with thrombin, the physiological fibrinogen-clotting enzyme, showed that thrombin’s fibrinogen-recognition exosite (FRE) is poorly represented in crotalase. Hirudin, a FRE-dependent inhibitor, had no effect on crotalase. Spatial modeling of crotalase yielded a possible alternative fibrinogen-recognition site comprised of Arg 60F, Lys 85, Lys 87, and Arg 107 (underlined in the sequence above). Crotalase also lacks thrombin’s YPPW loop, as well as its functionally important ETW 146-148, and its heparin-binding site. The enzyme contains a single asparagine-linked glycosylation site, NFT, bearing neutral and amino sugars that account for 8.3% of the enzyme’s total molecular weight of 29,027. The calculated absorbance of crotalase at 280 nm, 1%, cm-1is 15.2.

Die primäre Proteinstruktur von Stämmen des Tabakmosaikvirus

1963 ◽  
Vol 18 (12) ◽  
pp. 1032-1049 ◽  
Author(s):  
B. Wittmann-Liebold ◽  
H. G. Wittmann
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Amino Acid Sequences ◽  
Tryptic Peptides

The amino acid sequence of dahlemense, a naturally occuring strain of tobacco mosaic virus, has been determined and compared with that of the strain vulgare (Fig. 7). In this communication the experimental details are given for the elucidation of the amino acid sequences within two tryptic peptides with 65 amino acids.

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