Interaction of calpastatin with calpain: a review

Abstract

Calpastatin is a multiheaded inhibitor capable of inhibiting more than one calpain molecule. Each inhibitory domain of calpastatin has three subdomains, A, B, and C; A binds to domain IV and C binds to domain VI of the calpains. Crystallographic evidence shows that binding of C to domain VI involves hydrophobic interactions at a site near the first EF-hand in domain VI. Sequence homology suggests that binding of A to calpain domain IV also involves hydrophobic interactions near the EF1-hand of domain IV. Neither subdomain A nor C have inhibitory activity without subdomain B, but both increase the inhibitory activity of B. Subdomain B peptides have no inhibitory activity unless they contain at least 13 amino acids, and inhibitory activity increases with the number of amino acid residues, suggesting that inhibition requires interaction over a large area of the calpain molecule. Although subdomain B inhibition kinetically is competitive in nature, subdomain B does not seem to interact with the active site of the calpains directly, but may bind to domain III of the calpains and act to block access to the active site. It is possible that subdomain B binds to calpain only after it has been activated by Ca2+.

Calcium‐induced tripartite binding of intrinsically disordered calpastatin to its cognate enzyme, calpain

“…fast exchange was observed in the whole concentration range (data not shown), in accord with previous observations (reviewed in Refs. [1,3]). …”

“…For the first time, subdomains were defined as three well-conserved regions encoded by separate exons [26,29]. Other sequence alignment studies have suggested that A is located between S 12 and G 30 and C is between G 91 and T 104 [3]. These segments have been suggested as forming a-helices when bound to calpain, and interference with their helix-forming potential by point mutations does cause a drastic decrease in binding strength [30].…”

“…Every inhibitory domain contains three short conserved segments, termed subdomains, which are primarily responsible for calpastatin binding to calpain. Subdomain A and C (or A and C) potentiate inhibition, as they tether the inhibitor to the enzyme in a calcium-dependent manner, but do not inhibit enzyme activity on their own [3,7]. The middle segment, B, binds in the active-site region of the enzyme and carries inhibitory potential in itself.…”

Regulating cell migration: calpains make the cut

“…Similarly, regulation of protein-protein interactions changes the calcium requirements of calpains (Melloni et al, 2000a;Melloni et al, 2000b;Melloni et al, 1998;Melloni et al, 2000c;Michetti et al, 1991;Salamino et al, 1993), but their roles in activation are not clear. Finally, calpains are regulated by their best-known interacting partner, the endogenous calpain inhibitor calpastatin (Wendt et al, 2004). Although overexpression of calpastatin in cells can decrease calpain activity, escape from calpastatin is not Fig.…”

Inhibition of human μ-calpain by conformationally constrained calpastatin peptides

“…Besides the local calcium concentration, the endogenous protein-type inhibitor calpastatin plays an important role in the regulation of calpain activity (for a recent review see Wendt et al, 2004). Calpastatin is widely distributed in mammalian tissues.…”