Abstract

The importance of individual residues in the N-terminal region of cystatin B for proteinase inhibition was elucidated by measurements of the affinity and kinetics of binding of N-terminally truncated, recombinant variants of the bovine inhibitor to cysteine proteinases. Removal of Met-1 caused an 8- to 10-fold lower affinity for papain and cathepsin B, decreased the affinity also for cathepsin L but only minimally affected cathepsin H affinity. Additional truncation of Met-2 further weakened the binding to papain and cathepsin B by 40-70-fold, whereas the affinity for cathepsins L and H was essentially unaffected. Removal of Cys-3 had the most drastic effects on the interactions, resulting in a further affinity decrease of similar to 1500-fold for papain, similar to 700-fold for cathepsin L and similar to 15-fold for cathepsin H; the binding to cathepsin B could not be assessed. The binding kinetics could only be evaluated for papain and cathepsin H and showed that the reduced affinities for these enzymes were predominantly due to increased dissociation rate constants. These results demonstrate that the N-terminal region of cystatin B contributes appreciably to proteinase inhibition, in contrast to previous proposals. It is responsible for 12-40% of the total binding energy of the inhibitor to the proteinases investigated, being of least importance for cathepsin H binding. Cys-3 is the most important residue of the N-terminal region for inhibition of papain, cathepsin L and cathepsin H, the role of the other residues of this region varying with the target proteinase. (C) 2002 Elsevier Science B.V. All rights reserved

Published in

BBA - Proteins and Proteomics
2003, Volume: 1645, number: 1, pages: 105-112
Publisher: ELSEVIER SCIENCE BV

SLU Authors

• Björk, Ingemar

  • Department of Molecular Biosciences, Swedish University of Agricultural Sciences
    

Publication Identifiers

DOI: https://doi.org/10.1016/S1570-9639(02)00526-5

Permanent link to this page (URI)

https://res.slu.se/id/publ/1083