CRYSTAL-STRUCTURE OF CUTINASE COVALENTLY INHIBITED BY A TRIGLYCERIDE ANALOG

Cutinase from Fusarium solani is a lipolytic enzyme that hydrolyses tr iglycerides efficiently. All the inhibited forms of lipolytic enzymes described so far are based on the use of small organophosphate and org anophosphonate inhibitors, which bear little resemblance to a natural triglyceride substrate. In this article we describe the crystal struct ure of cutinase covalently inhibited by rbamoylglycero-3-O-p-nitrophen ylbutyl-phosphonate, a triglyceride analogue mimicking the first tetra hedral intermediate along the reaction pathway. The structure, which h as been solved at 2.3 Angstrom, reveals that in both the protein molec ules of the asymmetric unit the inhibitor is almost completely embedde d in the active site crevice. The overall shape of the inhibitor is th at of a fork: the two dibutyl-carbamoyl chains point towards the surfa ce of the protein, whereas the butyl chain bound to the phosphorous at om is roughly perpendicular to the sn-1 and sn-2 chains. The sn-3 chai n is accommodated in a rather small pocket at the bottom of the active site crevice, thus providing a structural explanation for the prefere nce of cutinase for short acyl chain substrates.