Structural analysis of cysteine protease ubiquitin carboxy-terminal hydrolase UCHL1 in complex with the suicide inhibitor Z-VAE(OMe)-FMK

Joseph R Chaney, Purdue University

Abstract

Ubiquitin carboxy-terminal hydrolase L1 (UCHL1) is a Parkinson's disease-associated cysteine hydrolase found abundantly and selectively expressed in neurons. The three dimensional structure of UCHL1 has previously been determined by x-ray diffraction at 2.4 Å resolution. The structure reveals that the catalytic residues (Cys 90, His 161, Asp 176) are misaligned, with nucleophilic cysteine (Cys 90) 7.7 Å away from the general base His 161, consistent with an inactive form of the enzyme. In this thesis, analysis of the co-crystal structure of UCHL1 bound to the suicide inhibitor, benzyloxycarbonyl-valine-alanine-glutamic-acid (OMe) fluoromethylketone (ZVAE(OMe)-FMK), is described. This structure reveals the mechanism of UCHL1 inactivation by the suicide inhibitor and allows a visualization of the intermolecular contacts between the enzyme and the inhibitor. Inspection of the co-crystal structure reveals that the Cys 90 of the enzyme is covalently linked to the inhibitor via a thioester bond. While the covalent bond allows tethering of the inhibitor to the active-site cysteine, a number of intermolecular interactions are observed between the inhibitor and the residues of the enzyme lining the S&feet; side of the active-site cleft. The structure also reveals that the Z-VAE(OMe)-FMK inhibitor is bound to the active site with the catalytic triad in an unproductive form, in which the general base histidine (His 161) is 8.0 Å away from the catalytic Cys 90, implying that the inhibitor may be expected to exhibit selectivity over other ubiquitin hydrolases, such as UCHL3, which have canonical catalytic triads.

Degree

M.S.

Advisors

Das, Purdue University.

Subject Area

Biochemistry

COinS