Structures of the platelet calcium- and integrin-binding protein and the ?IIb-integrin cytoplasmic domain suggest a mechanism for calcium-regulated recognition; homology modelling and NMR studies

Abstract

Calcium- and integrin-binding protein (CIB) binds to the 20-residue alphaIIb cytoplasmic domain of platelet alphaIIbbeta3 integrin. Amino acid sequence similarities with calmodulin (CaM) and calcineurin B (CnB) allowed the construction of homology-based models of calcium-saturated CIB as well as apo-CIB. In addition, the solution structure of the alphaIIb cytoplasmic domain in 45% aqueous trifluoroethanol was solved by conventional two-dimensional NMR methods. The models indicate that the N-terminal domain of CIB possesses a number of positively charged residues in its binding site that could interact with the acidic carboxy-terminal LEEDDEEGE sequence of alphaIIb. The C-terminal domain of CIB seems well-suited to bind the sequence WKVGFFKR, which forms a well-structured alpha helix; this is analogous to calmodulin and calcineurin B, which also bind alpha helices. Similarities between the C-terminal domains of CIB and calmodulin suggest that binding of CIB to the cytoplasmic domain of alphaIIb may be affected by fluctuations in the intracellular calcium concentration.

Transmembrane signal transduction of the α IIb β 3 integrin

“…This kink is not in agreement with electron microscopy studies and might be a crystallization artifact. The structure of the cytosolic domain of α IIb has been solved by NMR in 45% polytetrafluoroethylene (Hwang and Vogel 2000), as well as associated to a micelle (Vinogradova et al 2000). For our studies, we chose the structure of the cytoplasmic domain of α IIb associated to a micelle because here the structure‐inducing effect of the membrane is mimicked.…”

Calcium-binding calmyrin forms stable covalent dimers in vitro, but in vivo is found in monomeric form.

“…Homology-based models of calmyrin assumed Ca 2+ -dependent exposure of hydrophobic residues in the C-terminal domain of calmyrin (Hwang & Vogel, 2000). Such exposure is typical for calmodulin and other calcium sensor proteins enabling subsequent binding to α-helical structures of their cellular targets.…”

CIB1: a small protein with big ambitions

“…To gain insights into CIB1 function, the structure of CIB1 has been explored extensively by NMR, circular dichroism, X-ray crystallography, and sequence analyses (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12). CIB1 is a small, 22-kDa, 191-aa, intracellular Ca 2+ -binding protein that possesses an N-terminal myristoylation site (3,13).…”