TY - JOUR
T1 - Requirements of different subdomains of calpastain for calpain inhibition and for binding to calmodulin-like domains
AU - Ma, Hong
AU - Qiong Yang,, Hong
AU - Takano,, Emiko
AU - Joo Lee,, Woon
AU - Hatanaka, Masakazu
AU - Maki, Masatoshi
PY - 1993/5
Y1 - 1993/5
N2 - Calpain requires Ca2+ for both proteolysis of its substrates and interaction with its endogenous inhibitor, calpastatin. The mechanism of inhibition of calpain by calpastatin has remained unsolved, although Nishimura and Goll [J. BioL Chem. 266, 11842-11850 (1991)] reported that autolyzed calpain fragments containing calmodulin-like domains (CaMLDs) bound to an immobilized calpastatin column. We investigated the correlation between CaMLD-binding and calpain inhibition using immobilized columns of geneengineered CaMLDs derived from the human t-calpain large subunit and various recombinant calpastatin mutants. Among the four internally repetitive inhibitory domains of calpastatin, each having conserved regions A, B, and C, only domains 1 and 4 showed the binding activity. The region B deletion mutant of domain 1, retaining the CaMLD-binding ability, no longer had the calpain inhibition activity, and became susceptible to proteolysis. In contrast, a synthetic oligopeptide of region B with moderate calpain inhibition activity did not bind to the column. Domain 3 acquired the binding ability on substitution of region A with that of domain 1. These results suggest that calpain inhibition and binding to the CaMLDs are not correlated or mediated by different subdomains of calpastatin. 1993
AB - Calpain requires Ca2+ for both proteolysis of its substrates and interaction with its endogenous inhibitor, calpastatin. The mechanism of inhibition of calpain by calpastatin has remained unsolved, although Nishimura and Goll [J. BioL Chem. 266, 11842-11850 (1991)] reported that autolyzed calpain fragments containing calmodulin-like domains (CaMLDs) bound to an immobilized calpastatin column. We investigated the correlation between CaMLD-binding and calpain inhibition using immobilized columns of geneengineered CaMLDs derived from the human t-calpain large subunit and various recombinant calpastatin mutants. Among the four internally repetitive inhibitory domains of calpastatin, each having conserved regions A, B, and C, only domains 1 and 4 showed the binding activity. The region B deletion mutant of domain 1, retaining the CaMLD-binding ability, no longer had the calpain inhibition activity, and became susceptible to proteolysis. In contrast, a synthetic oligopeptide of region B with moderate calpain inhibition activity did not bind to the column. Domain 3 acquired the binding ability on substitution of region A with that of domain 1. These results suggest that calpain inhibition and binding to the CaMLDs are not correlated or mediated by different subdomains of calpastatin. 1993
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U2 - 10.1093/oxfordjournals.jbchem.a124088
DO - 10.1093/oxfordjournals.jbchem.a124088
M3 - Article
C2 - 8340353
AN - SCOPUS:0027252107
SN - 0021-924X
VL - 113
SP - 591
EP - 599
JO - Journal of Biochemistry
JF - Journal of Biochemistry
IS - 5
ER -