Structure of a Myristoyl-ACP-Specific Thioesterase from Vibrio harveyi

Abstract

The crystal structure of a myristoyl acyl carrier protein specific thioesterase (C14ACP-TE) from a bioluminescent bacterium, Vibrio harveyi, was solved by multiple isomorphous replacement methods and refined to an R factor of 22% at 2.1-A resolution. This is the first elucidation of a three-dimensional structure of a thioesterase. The overall tertiary architecture of the enzyme resembles closely the consensus fold of the rapidly expanding superfamily of alpha/beta hydrolases, although there is no detectable homology with any of its members at the amino acid sequence level. Particularly striking similarity exists between the C14ACP-TE structure and that of haloalkane dehalogenase from Xanthobacter autotrophicus. Contrary to the conclusions of earlier studies [Ferri, S. R., & Meighen, E. A. (1991) J. Biol. Chem. 266, 12852-12857] which implicated Ser77 in catalysis, the crystal structure of C14ACP-TE reveals a lipase-like catalytic triad made up of Ser114, His241, and Asp211. Surprisingly, the gamma-turn with Ser114 in a strained secondary conformation (phi = 53 degrees, psi = -127 degrees), characteristic 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 acids. Site-directed mutagenesis and radioactive labeling support the catalytic function of Ser114. Crystallographic analysis of the Ser77-->Gly mutant at 2.5-A resolution revealed no structural changes; in both cases the loop containing the residue in position 77 is disordered.(ABSTRACT TRUNCATED AT 250 WORDS)

The crystal structure of palmitoyl protein thioesterase 1 and the molecular basis of infantile neuronal ceroid lipofuscinosis

“…The two thioesterases share the core six-stranded ␤-sheet, although ACTE also contains the antiparallel strands ␤1 and ␤2 at the N-terminal end, as well as a longer C terminus, which contains an additional ␣-helix and ␤-strand. The insertion between ␤6 and ␤7 that forms the palmitate binding domain corresponds topologically to the smaller ''cap'' subdomain predicted form the myristoylbinding pocket in ACTE (12), but it is structurally quite different. PPT1 and ACTE have a common catalytic triad and acyl transfer mechanism, but differ in specificity.…”

“…The Vibrio harveyi myristoyl-acyl carrier protein thioesterase (ACTE) (12), an enzyme involved in bioluminescence, contains the ␣͞␤ hydrolase fold characteristic of microbial lipases. The second, 4-hydroxybenzoyl-CoA thioesterase from Pseudomonas sp.…”

Palmitoyl-acyl carrier protein (ACP) thioesterase and the evolutionary origin of plant acyl-ACP thioesterases.

“…Extensive searches in current sequence data bases failed to uncover any significant relationships of FatAlB with any other entry. Even the known non-plant acyl-ACP thioesterases (Lawson et al, 1994) appear to have a different origin. To date, therefore, no clues for the evolutionary source of Fat, which could have assisted with the evaluation of its original specificity, have been uncovered by this approach.…”

Methyl Phenylalkanoates as Substrates to Probe the Active Sites of Esterases

“…Structural studies and the use of specific inhibitors such as diisopropylfluorophosphate have indicated that a serine residue forms the catalytic centre in many functionally diverse hydrolytic enzymes such as proteases [ 11, triacylglycerol lipases [2-51, acetylcholinesterases [6-71, butyrylcholinesterases [8], thioesterases [9], cholesterol esterases [lo] and even a peroxidase [11]. In most esterases, the serine forms part of a catalytic triad comprising histidine, serine and a carboxylic acid, and it has been postulated that the triad functions to increase the nucleophilicity of catalytic serine residue by means of a chargerelay system [l].…”