Multiple proteolytic action of rat liver cathepsin B: Specificities and pH-dependences of the endo- and exopeptidase activities

Hironobu Koga, Hidenori Yamada, Yukio Nishimura, Keitaro Kato, Taiji Imoto

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Abstract

Dipeptidylcarboxypeptidase, endopeptidase, and carboxypeptidase activities of rat liver cathepsin B were investigated using soluble denatured protein substrates, reduced and S-(3-trimethylammonio)propylated proteins and their derivatives. It was found that the soluble denatured proteins were degraded mainly by the dipeptidylcarboxypeptidase activity and in a few cases by the endopeptidase and carboxypeptidase activities. The dipeptidylcarboxypeptidase activity showed broad substrate specificity with broad pH optimum at 4-6. A peptide having the α-carboxyl group amidated with methylamine could also be a good substrate for this activity. These results suggest that this activity is dependent not upon the dissociated α-carboxyl group at the P2' site but upon the hydrogen-bonding abilities of the α-imino moiety and the protonated or amidated α-carboxyl moiety at P2'. On the other hand, the endopeptidase and carboxypeptidase activities were observed in a few cases, suggesting that special amino acid sequences in the substrates are responsible for these activities. These activities showed sharp pH optima at 6 and seemed to prefer basic amino acid residues at P1 site. Therefore, we suppose that cathepsin B has a carboxyl group with a pKa, of about 5.5 at the S1 subsite which more effectively interacts with a positive charge at the P1 site of the substrate at pH 6 than at pH 5. Based on these results, a model of the binding subsites of this enzyme is proposed.

Original languageEnglish
Pages (from-to)179-188
Number of pages10
JournalJournal of Biochemistry
Volume110
Issue number2
DOIs
Publication statusPublished - Jan 1 1991

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All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

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