STRUCTURE OF A MYRISTOYL-ACP-SPECIFIC THIOESTERASE FROM VIBRIO-HARVEYI

The crystal structure of a myristoyl acyl carrier protein specific thi oesterase (C(14)ACP-TE) from a bioluminescent bacterium, Vibrio harvey i, was solved by multiple isomorphous replacement methods and refined to an R factor of 22% at 2.1-Angstrom resolution. This is the first el ucidation of a three-dimensional structure of a thioesterase. The over all tertiary architecture of the enzyme resembles closely the consensu s fold of the rapidly expanding superfamily of alp hydrolases, althoug h there is no detectable homology with any of its members at the amino acid sequence level. Particularly striking similarity exists between the C(14)ACP-TE structure and that of haloalkane dehalogenase from Xan thobacter autotrophicus. Contrary to the conclusions of earlier studie s [Ferri, S. R., and Meighen, E. A (1998) J. Biol. Chem. 266, 12852-12 857] which implicated Ser77 in catalysis, the crystal structure of C(1 4)ACP-TE reveals a lipase-like catalytic triad made up of Ser114, His2 41, and Asp211. Surprisingly, the gamma-turn with Ser114 in a strained secondary conformation (phi = 53 degrees, psi = -127 degrees), charac teristic of the so-called nucleophilic elbow, does not conform to the frequently invoked lipase/esterase consensus sequence (Gly-X-Ser-X-Gly ), as the positions of both glycines are occupied by larger amino acid s. Site-directed mutagenesis and radioactive labeling support the cata lytic function of Ser 114. Crystallographic analysis of the Ser77 --> Gly mutant at 2.5-Angstrom resolution revealed no structural changes; in both cases the loop containing the residue in position 77 is disord ered. The oxyanion hole does not appear to be fully formed in the nati ve enzyme, suggesting that its structural reorganization may occur upo n substrate binding, a mechanism reminiscent of the conformational cha nges underlying interfacial activation in triglyceride lipases.