Structure and function relationship of staphylocoagulase

Shun-Ichiro Kawabata, Takashi Morita, Toshiyuki Miyata, Shigenori Kaida, Sadaaki Iwanaga, Hideo Igarashi

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

The bacterial protein staphylocoagulase binds stoichiometrically to human prothrombin, resulting in a coagulant complex, staphylothrombin. The enzymatic properties of staphylothrombin differ from those of α-thrombin in their substrate specificities toward natural and synthetic substrates, in addition to their interaction with protease inhibitors. In order to obtain information about the region of staphylocoagulase that interacts with human prothrombin, staphylocoagulase was cleaved by α-chymotrypsin. Limited α-chymotryptic cleavage of staphylocoagulase yielded three large fragments, of 43, 30, and 20 kD. The 43-kD fragment exhibited a high affinity for human prothrombin (Kd=1.7 nM), which is comparable to the affinity observed using intact staphylocoagulase (Kd=0.46 nM). A complex of the 43-kD fragment and prothrombin possessed both clotting and amidase activity essentially identical to that observed in a complex of intact staphylocoagulase and prothrombin. The 30-kD fragment exhibited weaker affinity for prothrombin (Kd=120 nM.) While clotting activity was not observed with a complex of this fragment and prothrombin, it nonetheless possessed a weak amidase activity. The 20-kD fragment was found only to bind to prothrombin. The NH2-terminal sequence analyses of these fragments revealed that the 43-kD fragment constitutes the NH2-terminal portion of staphylocoagulase, and contains the 30-kD and 20-kD fragments. It is therefore concluded that the functional region of staphylocoagulase for binding and activation of prothrombin is localized in the NH2-terminal region of the intact protein. The 43-kD fragment contained 324 amino acids with a molecular weight of 38,098. The 43-kD fragment had an unusual amino acid composition based on a sequence in which the sum of Asp (28 residues), Asn (22), Glu (35), Gln (9), and Lys (52) residues accounted for more than 45% of the total. A comparison of the amino acid sequence of the 43-kD fragment with that of streptokinase did not reveal any obvious sequence homology. There was also no sequence homology with that of trypsin, α-chymotrypsin, and elastase.

Original languageEnglish
Pages (from-to)17-32
Number of pages16
JournalJournal of Protein Chemistry
Volume6
Issue number1
DOIs
Publication statusPublished - Feb 1 1987

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Coagulase
Prothrombin
Amino acids
amidase
Substrates
Chemical activation
Molecular weight
Sequence Homology
Proteins
Amino Acids
Chemical analysis
Coagulants
Streptokinase
Bacterial Proteins
Pancreatic Elastase
Chymotrypsin
Substrate Specificity
Protease Inhibitors
Thrombin
Sequence Analysis

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Kawabata, S-I., Morita, T., Miyata, T., Kaida, S., Iwanaga, S., & Igarashi, H. (1987). Structure and function relationship of staphylocoagulase. Journal of Protein Chemistry, 6(1), 17-32. https://doi.org/10.1007/BF00248824

Structure and function relationship of staphylocoagulase. / Kawabata, Shun-Ichiro; Morita, Takashi; Miyata, Toshiyuki; Kaida, Shigenori; Iwanaga, Sadaaki; Igarashi, Hideo.

In: Journal of Protein Chemistry, Vol. 6, No. 1, 01.02.1987, p. 17-32.

Research output: Contribution to journalArticle

Kawabata, S-I, Morita, T, Miyata, T, Kaida, S, Iwanaga, S & Igarashi, H 1987, 'Structure and function relationship of staphylocoagulase', Journal of Protein Chemistry, vol. 6, no. 1, pp. 17-32. https://doi.org/10.1007/BF00248824
Kawabata, Shun-Ichiro ; Morita, Takashi ; Miyata, Toshiyuki ; Kaida, Shigenori ; Iwanaga, Sadaaki ; Igarashi, Hideo. / Structure and function relationship of staphylocoagulase. In: Journal of Protein Chemistry. 1987 ; Vol. 6, No. 1. pp. 17-32.
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abstract = "The bacterial protein staphylocoagulase binds stoichiometrically to human prothrombin, resulting in a coagulant complex, staphylothrombin. The enzymatic properties of staphylothrombin differ from those of α-thrombin in their substrate specificities toward natural and synthetic substrates, in addition to their interaction with protease inhibitors. In order to obtain information about the region of staphylocoagulase that interacts with human prothrombin, staphylocoagulase was cleaved by α-chymotrypsin. Limited α-chymotryptic cleavage of staphylocoagulase yielded three large fragments, of 43, 30, and 20 kD. The 43-kD fragment exhibited a high affinity for human prothrombin (Kd=1.7 nM), which is comparable to the affinity observed using intact staphylocoagulase (Kd=0.46 nM). A complex of the 43-kD fragment and prothrombin possessed both clotting and amidase activity essentially identical to that observed in a complex of intact staphylocoagulase and prothrombin. The 30-kD fragment exhibited weaker affinity for prothrombin (Kd=120 nM.) While clotting activity was not observed with a complex of this fragment and prothrombin, it nonetheless possessed a weak amidase activity. The 20-kD fragment was found only to bind to prothrombin. The NH2-terminal sequence analyses of these fragments revealed that the 43-kD fragment constitutes the NH2-terminal portion of staphylocoagulase, and contains the 30-kD and 20-kD fragments. It is therefore concluded that the functional region of staphylocoagulase for binding and activation of prothrombin is localized in the NH2-terminal region of the intact protein. The 43-kD fragment contained 324 amino acids with a molecular weight of 38,098. The 43-kD fragment had an unusual amino acid composition based on a sequence in which the sum of Asp (28 residues), Asn (22), Glu (35), Gln (9), and Lys (52) residues accounted for more than 45{\%} of the total. A comparison of the amino acid sequence of the 43-kD fragment with that of streptokinase did not reveal any obvious sequence homology. There was also no sequence homology with that of trypsin, α-chymotrypsin, and elastase.",
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