The crystal and molecular structure of human annexin V, an anticoagulant protein that binds to calcium and membranes.

Abstract

Human annexin V (PP4), a member of the family of calcium, membrane binding proteins, has been crystallized in the presence of calcium and analysed by crystallography by multiple isomorphic replacement at 3 A and preliminarily refined at 2.5 A resolution. The molecule has dimensions of 64 x 40 x 30 A3 and is folded into four domains of similar structure. Each domain consists of five alpha‐helices wound into a right‐handed superhelix yielding a globular structure of approximately 18 A diameter. The domains have hydrophobic cores whose amino acid sequences are conserved between the domains and within the annexin family of proteins. The four domains are folded into an almost planar array by tight (hydrophobic) pair‐wise packing of domains II and III and I and IV to generate modules (II‐III) and (I‐IV), respectively. The assembly is symmetric with three parallel approximate diads relating II to III, I to IV and the module (II‐III) to (I‐IV), respectively. The latter diad marks a channel through the centre of the molecule coated with charged amino acid residues. The protein has structural features of channel forming membrane proteins and a polar surface characteristic of soluble proteins. It is a member of the third class of amphipathic proteins different from soluble and membrane proteins.

Mapping of three unique Ca2+‐binding sites in human annexin II

“…the residues directly preceding Gly206, is the only bond in the annexin I1 core which is susceptible to limited trypsin digestion, indicating that this region is exposed at the surface of the protein (Johnson and Weber, 1990). In addition, the corresponding region in each repeat of annexin V forms an interhelical loop as determined by X-ray crystallography (Huber et al, 1990a).…”

“…The recently determined crystal structure of one member of the annexin family, annexin V, revealed that the protein core forms a highly symmetric unit with densely packed CIhelices (Huber et al, 1990a). Each annexin repeat is composed of five a-helices (a-e) which are connected via short loops or turns (Huber et al, 1990a).…”

“…Each annexin repeat is composed of five a-helices (a-e) which are connected via short loops or turns (Huber et al, 1990a). In the individual repeat the endonexin fold covers helix b and the preceding interhelical loop between helices a and b which contains the highly conserved glycine and threonine residues (positions 4 and 5 of the endonexin fold).…”

“…phospholipid binding is only observed in the presence of Ca2+, and the affinity for Ca2+ is increased by at least two orders of magnitude in the presence of phospholipid (for review see Klee, 1988). In the annexin V crystals, all Ca2+ sites are located on one side of the molecule, which exhibits a more convex shape, while the N-terminus, which is relatively short in annexin V, resides on the opposite concave surface (Huber et al, 1990a).…”

Mechanism of Annexin I-Mediated Membrane Aggregation

“…Despite fundamental differences, these models share a common concept of two distinct interaction sites; a primary site for membrane binding and a secondary site for membrane aggregation. The primary membranebinding site, which has been defined by extensive structural and mutational studies, coincides with the calcium-binding sites located on the convex surface of annexin molecules (20)(21)(22)(23)(24)(25)(26)(27). Much less is known about the secondary interaction site, although it has been postulated to be on the opposite concave side of the annexin molecule that harbors the aminoterminal region (19,28,29).…”