Refined 2.0 A X-ray crystal structure of the snake venom zinc-endopeptidase adamalysin II. Primary and tertiary structure determination, refinement, molecular structure and comparison with astacin, collagenase and thermolysin - PubMed (original) (raw)

Comparative Study

. 1994 Jun 17;239(4):513-44.

doi: 10.1006/jmbi.1994.1392.

Affiliations

Comparative Study

Refined 2.0 A X-ray crystal structure of the snake venom zinc-endopeptidase adamalysin II. Primary and tertiary structure determination, refinement, molecular structure and comparison with astacin, collagenase and thermolysin

F X Gomis-Rüth et al. J Mol Biol. 1994.

Abstract

Adamalysin II, alias proteinase II, a 24 kDa zinc-endopeptidase isolated from the snake venom of the Eastern diamondback rattlesnake Crotalus adamanteus, is a prototype of the proteolytic domain of snake venom metalloproteinases and of domains found in mammalian reproductive tract proteins. Its 2.0 A crystal and molecular structure was solved by multiple isomorphous replacement using six heavy-atom derivatives, and was refined to a crystallographic R-value of 0.172. 201 of the 203 amino acid residues of adamalysin II are defined by electron density; only the first two residues are disordered and crystallographically undefined in the crystal structure. Three-quarters of these crystallographic amino acid residue assignments were confirmed by chemical sequencing. In addition, the active-site zinc-ion, a hepta-coordinated calcium ion, a fixed sulphate anion and 173 solvent molecules were localized in the structure. Adamalysin II is an ellipsoidal molecule with a relatively flat active-site cleft separating the "upper" main body from a small "lower" subdomain. The regularly folded N-terminal upper domain consists essentially of a central, highly twisted five-stranded beta-pleated sheet flanked by a long and a short surface located helix on its convex side, and by two long helices, one of which represents the central "active site helix", on its concave side. The lower subdomain, comprising the last 50 residues, is organized in multiple turns, with the chain ending in a long C-terminal helix and an extended segment clamped to the upper domain via a disulphide bridge. The catalytic zinc-ion, located at the bottom of the active-site cleft, is almost tetrahedrally co-ordinated by His142, His146 and His152, and a water molecule anchored to an intermediate glutamic acid residue (Glu143), with the three imidazole N epsilon 2 nitrogen atoms 2.1 A and the solvent oxygen atom 2.4 A away from the zinc ion. His142, Glu143 and His146 are part of the long active-site helix, which extends up to Gly149, where it turns sharply away towards His152. The importance of these residues for structure and activity of adamalysin II explains their occurrence in the HEXXHXXGXXH consensus sequence. Asp153, which is strictly conserved in these snake venom and reproductive tract metalloproteinases, is buried in the subdomain and seems to stabilize the hydrophobic active-site basement. Some residues behind, the adamalysin peptide chain folds into a characteristic 1,4-turn (the "Met-turn") containing the conserved Met166, which forms a hydrophobic basement for the three zinc-binding imidazoles.(ABSTRACT TRUNCATED AT 400 WORDS)

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