Thioesterases: A new perspective based on their primary and tertiary structures

Abstract

Thioesterases (TEs) are classified into EC 3.1.2.1 through EC 3.1.2.27 based on their activities on different substrates, with many remaining unclassified (EC 3.1.2.-). Analysis of primary and tertiary structures of known TEs casts a new light on this enzyme group. We used strong primary sequence conservation based on experimentally proved proteins as the main criterion, followed by verification with tertiary structure superpositions, mechanisms, and catalytic residue positions, to accurately define TE families. At present, TEs fall into 23 families almost completely unrelated to each other by primary structure. It is assumed that all members of the same family have essentially the same tertiary structure; however, TEs in different families can have markedly different folds and mechanisms. Conversely, the latter sometimes have very similar tertiary structures and catalytic mechanisms despite being only slightly or not at all related by primary structure, indicating that they have common distant ancestors and can be grouped into clans. At present, four clans encompass 12 TE families. The new constantly updated ThYme (Thioesteractive enzYmes) database contains TE primary and tertiary structures, classified into families and clans that are different from those currently found in the literature or in other databases. We review all types of TEs, including those cleaving CoA, ACP, glutathione, and other protein molecules, and we discuss their structures, functions, and mechanisms.

α-Carbonic Anhydrases Possess Thioesterase Activity

“…26 For 15 out of 27 such groups, the substrates are thioesters of coenzyme A (CoA), whereas two contain acyl carrier proteins (ACP), four have glutathione or its derivatives as substrates, one has ubiquitin, and two such enzyme families possess other diverse substrates. 26,27 One important aspect of this enzyme superfamily is that they are not metalloenzymes except for one family, the hydroxyglutathione hydrolases (glyoxalases II), which are zinc enzymes with a metallo-β-lactamase fold. 26 Indeed, in all other thioesterase clans known to date the thioester hydrolysis is achieved by a catalytic dyad/triad normally comprising a nucleophilic amino acid−histidine− acidic amino acid sequence.…”

Functional Screening and In Vitro Analysis Reveal Thioesterases with Enhanced Substrate Specificity Profiles That Improve Short-Chain Fatty Acid Production in Escherichia coli

“…Selecting individual thioesterase enzymes for functional screening against short-chain fatty acyl-CoAs is challenging because much of their vast phylogenetic and functional diversity is poorly understood (13). While many thioesterases have been ex-plored for long-chain fatty acid production (14)(15)(16)(17), few studies have focused on those that prefer short-chain acyl-CoAs.…”

Acyl carrier protein structural classification and normal mode analysis

“…16 There we have classified ACPs into families, following the same techniques that we have used earlier with thioesterases 17 and ketoacyl synthases, 18 which are described in Computational Methods. In general, members of a protein family have strong sequence similarity.…”

“…[16][17][18] In brief, families were based on sequences with evidence at protein level from the UniProt database 53 to query the nonredundant (nr) protein sequence database. 55 A cutoff E-value of 0.001 (likelihood that similarity between query and compared sequence is due to chance) was used as the exclusion criterion.…”

“…60 All RMSD values were calculated between a-carbon atoms using MATLAB, to consider the most possible a-carbon atoms in the calculation; MultiProt reports the RMSD for only aligned residues. Values of RMSD (between two structures), RMSD ave (between three or more structures), and P ave (the average percentage of a-carbon atoms of the amino acid residues used to calculate the RMSD between two compared structures) were calculated, as explained in detail in the Supporting Information in Cantu et al 17 When a NMR-resolved structure was superimposed, only the lowest-energy frame was used, chosen following single-point energy calculations with the ff99SB AMBER force field. 61 …”