Adenylate-forming enzymes

Abstract

Thioesters, amides, and esters are common chemical building blocks in a wide array of natural products. The formation of these bonds can be catalyzed in a variety of ways. For chemists, the use of an activating group is a common strategy and adenylate enzymes are exemplars of this approach. Adenylating enzymes activate the otherwise unreactive carboxylic acid by transforming the normal hydroxyl leaving group into adenosine monophosphate. Recently there have been a number of studies of such enzymes and in this review we suggest a new classification scheme. The review highlights the diversity in enzyme fold, active site architecture, and metal coordination that has evolved to catalyze this particular reaction.

Figure 1

a Schematic representation of the two step process of adenylation. In the first reaction step ATP (A) and substrate are bound and the reactive substrate-adenylate and pyrophosphate (PP_i) formed. In some cases PP_i is released at this stage. In the second step a nucleophile (N) reacts with the reactive intermediate and the final product and adenosine monophosphate are released. In some cases PP_i is released at this stage. b Old and new classification of adenylate-forming enzymes. c Chemical representation of the two step adenylation reaction by adenylate-forming enzymes to carboxylate an alcohol, amid or sulfhydryl moiety of the nucleophile.

Figure 2

Members of the different classes of adenylate-forming enzymes, N- and C-terminus are marked with blue and red spheres, respectively. The N-terminal and C-terminal domain are colored in green and yellow, respectively a class I: non-ribosomal independent Gramicidin synthetase N-terminal adenylation domain in complex with AMP and phenylalanine (PDB code: 1amu) AMP white carbons, phenylalanine grey carbons; b class I: crystal structure of a predicated acyl-CoA synthetase from E.coli (3dmy); c class II: LeuRS form T. thermophilus (1obc) a subclass IIa aminoacyl tRNA synthetase; d class II: monomer of human GlyRS (2pme) a subclass IIb aminoacyl tRNA synthetase; e class III Achromobactin synthetase protein D (AcsD) in complex with ATP, monomer (PDB code: 2w02)

Figure 3

Active site alignment of different classes of adenylate-forming enzymes. Alignment was carried out on α-phosphate of ATP or AMP. a Alignment of class I enzymes of Acetyl-CoA synthetase (2p2f), gramicidin S synthetase 1 (1amu), Japanese firefly luciferase (2d1q) and 4-Chlorobenzoyl-CoA ligase (3cw8). b Alignment of class II enzymes tryptophanyl-tRNA synthetase (1m83), human tryptophanyl-tRNA synthetase (2qui) and lysyl-tRNA synthetase (1e24), alignment of α-phosphate of ATP is shown, ATP in grey line representation is ATP of 2qui. c Alignment of class III enzyme: AcsD colored in blue (2w02) and AlcC colored in green, Mg ion in AcsD and AlcC show an α-γ phosphate coordination, while AlcC coordinates a second Mg ion in the α-β position (green). In contrast to class I and II water in class III enzymes is close to the α phosphate.

Figure 4

Coordination of α-phosphate by positively charged residues in different classes of adenylate-forming enzymes. In each member at least one positively charged residue, as lysine, argentine or histidine is present. Acetyl-CoA synthetase (2p2f) and Japanese firefly luciferase (2d1q) are class I, human tryptophanyl-tRNA synthetase (2qui) and lysyl-tRNA synthetase (1e24) are class II, AcsD (2wo2) and AlcC (alcc) are class III adenylate-forming enzymes. ATP colored in grey and shown in line representation is ATP of AcsD.