Identification of metal ligands in the Clostridium histolyticum ColH collagenase

Chang Min Jung, Osamu Matsushita, Seiichi Katayama, Junzaburo Minami, Jun Sakurai, Akinobu Okabe

Research output: Contribution to journalArticle

67 Citations (Scopus)

Abstract

A Clostridium histolyticum 116-kDa collagenase has an H415EXXH motif but not the third zinc ligand, as found in already characterized zinc metalloproteinases. To identify its catalytic site, we mutated the codons corresponding to the three conserved residues in the motif to other amino acid residues. The mutation affecting His415 or His419 abolished catalytic activity and zinc binding, while that affecting Glu416 did the former but not the latter. These results suggest that the motif forms the catalytic site. We also mutated the codons corresponding to other amino acid residues that are likely zinc ligands. The mutation affecting Glu447 decreased markedly both the enzymatic activity and the zinc content, while that affecting Glu446 or Glu451 had smaller effects on activity and zinc binding. These mutations caused a decrease in k(cat) but no significant change in K(m). These results are consistent with the hypothesis that Glu447 is the third zinc ligand. The spacing of the three zinc ligands is the same in all known clostridial collagenases but not in other known gluzincins, indicating that they form a new gluzincin subfamily. The effects of mutations affecting Glu446 and Glu451 suggest that the two residues are also involved in catalysis, possibly through an interaction with the two zinc-binding histidine residues.

Original languageEnglish
Pages (from-to)2816-2822
Number of pages7
JournalJournal of Bacteriology
Volume181
Issue number9
Publication statusPublished - 1999
Externally publishedYes

Fingerprint

Microbial Collagenase
Zinc
Metals
Ligands
Mutation
Collagenases
Codon
Clostridium histolyticum
Catalytic Domain
Amino Acids
Metalloproteases
Catalysis
Histidine

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Immunology

Cite this

Jung, C. M., Matsushita, O., Katayama, S., Minami, J., Sakurai, J., & Okabe, A. (1999). Identification of metal ligands in the Clostridium histolyticum ColH collagenase. Journal of Bacteriology, 181(9), 2816-2822.

Identification of metal ligands in the Clostridium histolyticum ColH collagenase. / Jung, Chang Min; Matsushita, Osamu; Katayama, Seiichi; Minami, Junzaburo; Sakurai, Jun; Okabe, Akinobu.

In: Journal of Bacteriology, Vol. 181, No. 9, 1999, p. 2816-2822.

Research output: Contribution to journalArticle

Jung, CM, Matsushita, O, Katayama, S, Minami, J, Sakurai, J & Okabe, A 1999, 'Identification of metal ligands in the Clostridium histolyticum ColH collagenase', Journal of Bacteriology, vol. 181, no. 9, pp. 2816-2822.
Jung, Chang Min ; Matsushita, Osamu ; Katayama, Seiichi ; Minami, Junzaburo ; Sakurai, Jun ; Okabe, Akinobu. / Identification of metal ligands in the Clostridium histolyticum ColH collagenase. In: Journal of Bacteriology. 1999 ; Vol. 181, No. 9. pp. 2816-2822.
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