Amino-terminal conserved region in proteinase inhibitor domain of calpastatin potentiates its calpain inhibitory activity by interacting with calmodulin-like domain of the proteinase

Hong Ma, Hong Qiong Yang, Emiko Takano, Masakazu Hatanaka, Masatoshi Maki

Research output: Contribution to journalArticle

61 Citations (Scopus)

Abstract

Calpastatin is a widely distributed endogenous inhibitor protein specifically acting on calpain (Ca2+-dependent proteinase) and is known to interact with the calmodulin-like domain (CaMLD) of the proteinase in a Ca2+-dependent fashion. The calpastatin molecule consists of four inhibitory domains (domains 1-4) with mutually homologous sequences in three regions designated as A, B, and C. Acidic amphiphilic α-helical motifs are found in both regions A and C. We investigated the correlation between the calpain inhibition potency and the ability of calpastatin to bind to recombinant CaMLD of the μ-calpain large subunit using various mutant proteins of pig calpastatin domain 1 expressed in Escherichia coli. Substitution of conserved Leu-161 with Pro in region A caused a reduction in activity of both calpain inhibition and CaMLD binding. Additional substitution of Leu-236 with Pro in region C further decreased the calpain inhibitory activity and caused a loss of CaMLD binding ability. The effects of mutation in region C alone on the above activities were smaller than those in region A. Although a mutant of deletion in the entire region B had no calpain inhibitory activity, it retained the CaMLD binding ability. On the other hand, although a region B oligopeptide had a moderate inhibitory activity, it had no CaMLD binding ability. These results suggest that region A has a role in potentiating the inhibitory activity of calpastatin by interacting with CaMLD of calpain to form a tighter complex where region B exerts the inhibitory function.

Original languageEnglish (US)
Pages (from-to)24430-24436
Number of pages7
JournalJournal of Biological Chemistry
Volume269
Issue number39
StatePublished - Sep 30 1994
Externally publishedYes

Fingerprint

Calpain
Calmodulin
Peptide Hydrolases
Substitution reactions
Oligopeptides
Mutant Proteins
Sequence Homology
calpastatin
Escherichia coli
Swine
Mutation
Molecules

ASJC Scopus subject areas

  • Biochemistry

Cite this

Amino-terminal conserved region in proteinase inhibitor domain of calpastatin potentiates its calpain inhibitory activity by interacting with calmodulin-like domain of the proteinase. / Ma, Hong; Yang, Hong Qiong; Takano, Emiko; Hatanaka, Masakazu; Maki, Masatoshi.

In: Journal of Biological Chemistry, Vol. 269, No. 39, 30.09.1994, p. 24430-24436.

Research output: Contribution to journalArticle

@article{ee8ddeec337c4e78b90ef895adfa365a,
title = "Amino-terminal conserved region in proteinase inhibitor domain of calpastatin potentiates its calpain inhibitory activity by interacting with calmodulin-like domain of the proteinase",
abstract = "Calpastatin is a widely distributed endogenous inhibitor protein specifically acting on calpain (Ca2+-dependent proteinase) and is known to interact with the calmodulin-like domain (CaMLD) of the proteinase in a Ca2+-dependent fashion. The calpastatin molecule consists of four inhibitory domains (domains 1-4) with mutually homologous sequences in three regions designated as A, B, and C. Acidic amphiphilic α-helical motifs are found in both regions A and C. We investigated the correlation between the calpain inhibition potency and the ability of calpastatin to bind to recombinant CaMLD of the μ-calpain large subunit using various mutant proteins of pig calpastatin domain 1 expressed in Escherichia coli. Substitution of conserved Leu-161 with Pro in region A caused a reduction in activity of both calpain inhibition and CaMLD binding. Additional substitution of Leu-236 with Pro in region C further decreased the calpain inhibitory activity and caused a loss of CaMLD binding ability. The effects of mutation in region C alone on the above activities were smaller than those in region A. Although a mutant of deletion in the entire region B had no calpain inhibitory activity, it retained the CaMLD binding ability. On the other hand, although a region B oligopeptide had a moderate inhibitory activity, it had no CaMLD binding ability. These results suggest that region A has a role in potentiating the inhibitory activity of calpastatin by interacting with CaMLD of calpain to form a tighter complex where region B exerts the inhibitory function.",
author = "Hong Ma and Yang, {Hong Qiong} and Emiko Takano and Masakazu Hatanaka and Masatoshi Maki",
year = "1994",
month = "9",
day = "30",
language = "English (US)",
volume = "269",
pages = "24430--24436",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "39",

}

TY - JOUR

T1 - Amino-terminal conserved region in proteinase inhibitor domain of calpastatin potentiates its calpain inhibitory activity by interacting with calmodulin-like domain of the proteinase

AU - Ma, Hong

AU - Yang, Hong Qiong

AU - Takano, Emiko

AU - Hatanaka, Masakazu

AU - Maki, Masatoshi

PY - 1994/9/30

Y1 - 1994/9/30

N2 - Calpastatin is a widely distributed endogenous inhibitor protein specifically acting on calpain (Ca2+-dependent proteinase) and is known to interact with the calmodulin-like domain (CaMLD) of the proteinase in a Ca2+-dependent fashion. The calpastatin molecule consists of four inhibitory domains (domains 1-4) with mutually homologous sequences in three regions designated as A, B, and C. Acidic amphiphilic α-helical motifs are found in both regions A and C. We investigated the correlation between the calpain inhibition potency and the ability of calpastatin to bind to recombinant CaMLD of the μ-calpain large subunit using various mutant proteins of pig calpastatin domain 1 expressed in Escherichia coli. Substitution of conserved Leu-161 with Pro in region A caused a reduction in activity of both calpain inhibition and CaMLD binding. Additional substitution of Leu-236 with Pro in region C further decreased the calpain inhibitory activity and caused a loss of CaMLD binding ability. The effects of mutation in region C alone on the above activities were smaller than those in region A. Although a mutant of deletion in the entire region B had no calpain inhibitory activity, it retained the CaMLD binding ability. On the other hand, although a region B oligopeptide had a moderate inhibitory activity, it had no CaMLD binding ability. These results suggest that region A has a role in potentiating the inhibitory activity of calpastatin by interacting with CaMLD of calpain to form a tighter complex where region B exerts the inhibitory function.

AB - Calpastatin is a widely distributed endogenous inhibitor protein specifically acting on calpain (Ca2+-dependent proteinase) and is known to interact with the calmodulin-like domain (CaMLD) of the proteinase in a Ca2+-dependent fashion. The calpastatin molecule consists of four inhibitory domains (domains 1-4) with mutually homologous sequences in three regions designated as A, B, and C. Acidic amphiphilic α-helical motifs are found in both regions A and C. We investigated the correlation between the calpain inhibition potency and the ability of calpastatin to bind to recombinant CaMLD of the μ-calpain large subunit using various mutant proteins of pig calpastatin domain 1 expressed in Escherichia coli. Substitution of conserved Leu-161 with Pro in region A caused a reduction in activity of both calpain inhibition and CaMLD binding. Additional substitution of Leu-236 with Pro in region C further decreased the calpain inhibitory activity and caused a loss of CaMLD binding ability. The effects of mutation in region C alone on the above activities were smaller than those in region A. Although a mutant of deletion in the entire region B had no calpain inhibitory activity, it retained the CaMLD binding ability. On the other hand, although a region B oligopeptide had a moderate inhibitory activity, it had no CaMLD binding ability. These results suggest that region A has a role in potentiating the inhibitory activity of calpastatin by interacting with CaMLD of calpain to form a tighter complex where region B exerts the inhibitory function.

UR - http://www.scopus.com/inward/record.url?scp=0028148057&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028148057&partnerID=8YFLogxK

M3 - Article

C2 - 7929105

AN - SCOPUS:0028148057

VL - 269

SP - 24430

EP - 24436

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 39

ER -