Structure of astacin and implications for activation of astacins and zinc-ligation of collagenases

Abstract

ASTACIN, a digestive zinc-endopeptidase from the crayfish Astacus astacus L.1,2, is the prototype for the 'astacin family'3–5, which includes mammalian metallo-endopeptidases5 and develop-mentally regulated proteins of man6, fruitfly7, frog8 and sea urchin9,10. Here we report the X-ray crystal structure of astacin, which reveals a deep active-site cleft, with the zinc at its bottom ligated by three histidines, a water molecule and a more remote tyrosine. The third histidine (His 102) forms part of a consensus sequence, shared not only by the members of the astacin family, but also by otherwise sequentially unrelated proteinases, such as vertebrate collagenases11. It may therefore represent the elusive 'third' zinc ligand in these enzymes. The amino terminus of astacin is buried forming an internal salt-bridge with Glu 103, adjacent to His 102. Astacin pro-forms extended at the N terminus, as observed for some 'latent' mammalian astacin homologues, did not exhibit this 'active' conformation, indicating an activation mechanism reminiscent of trypsin-like serine proteinases.