Crystal structures of the psychrophilic α-amylase from Alteromonas haloplanctis in its native form and complexed with an inhibitor

Abstract

Alteromonas haloplanctis is a bacterium that flourishes in Antarctic sea-water and it is considered as an extreme psychrophile. We have determined the crystal structures of the α-amylase (AHA) secreted by this bacterium, in its native state to 2.0 Å resolution as well as in complex with Tris to 1.85 Å, resolution. The structure of AHA, which is the first experimentally determined three-dimensional structure of a psychrophilic enzyme, resembles those of other known α-amylases of various origins with a surprisingly greatest similarity to mammalian α-amylases. AHA contains a chloride ion which activates the hydrolytic cleavage of substrate α-1,4-glycosidic bonds. The chloride binding site is situated ~5 Å from the active site which is characterized by a triad of acid residues (Asp 174, Glu 200, Asp 264). These are all involved in firm binding of the Tris moiety. A reaction mechanism for substrate hydrolysis is proposed on the basis of the Tris inhibitor binding and the chloride activation. A trio of residues (Ser 303, His 337, Glu 19) having a striking spatial resemblance with serine-protease like catalytic triads was found ~22 Å, from the active site. We found that this triad is equally present in other chloride dependent α-amylases, and suggest that it could be responsible for autoproleolytic events observed in solution for his cold adapted α-amylase.