title: Insights into the biosynthesis of the Vibrio cholerae major autoinducer CAI-1 from the crystal structure of the PLP-dependent enzyme CqsA.
abstract: CqsA is an enzyme involved in the biosynthesis of cholerae autoinducer-1 (CAI-1), the major Vibrio cholerae autoinducer engaged in quorum sensing. The amino acid sequence of CqsA suggests that it belongs to the family of alpha-oxoamine synthases that catalyse the condensation of an amino acid to an acyl-CoA substrate. Here we present the apo- and PLP-bound crystal structures of CqsA and confirm that it shares structural homology with the dimeric alpha-oxoamine synthases, including a conserved PLP-binding site. The chemical structure of CAI-1 suggests that decanoyl-CoA may be one substrate of CqsA and that another substrate may be l-threonine or l-2-aminobutyric acid. A crystal structure of CqsA at 1.9-A resolution obtained in the presence of PLP and l-threonine reveals an external aldimine that has lost the l-threonine side chain. Similarly, a 1.9-A-resolution crystal structure of CqsA in the presence of PLP, l-threonine, and decanoyl-CoA shows a trapped external aldimine intermediate, suggesting that the condensation and decarboxylation steps have occurred, again with loss of the l-threonine side chain. It is suggested that this side-chain loss, an observation supported by mass spectrometry, is due to a retro-aldol reaction. Although no structural data have been obtained on CqsA using l-2-aminobutyric acid and decanoyl-CoA as substrates, mass spectrometry confirms the expected product of the enzyme reaction. It is proposed that a region of structure that is disordered in the apo structure is involved in the release of product. While not confirming if CqsA alone is able to synthesize CAI-1, these results suggest possible synthetic routes.
authors.affiliation: Centre for Biomolecular Sciences, University of St. Andrews, UK.
authors.forename: Nasrin
authors.initials: N
authors.lastname: Jahan
authors.forename: Jane A
authors.initials: JA
authors.lastname: Potter
authors.forename: Md Arif
authors.initials: MA
authors.lastname: Sheikh
authors.forename: Catherine H
authors.initials: CH
authors.lastname: Botting
authors.forename: Sally L
authors.initials: SL
authors.lastname: Shirran
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Westwood
authors.forename: Garry L
authors.initials: GL
authors.lastname: Taylor
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D018360:Crystallography, X-Ray; D006801:Humans; D007659:Ketones / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D011487:Protein Conformation; D012545:Schiff Bases / Q000737:chemistry; D016415:Sequence Alignment; D014734:Vibrio cholerae / Q000201:enzymology*
chemical_list: C525620:3-hydroxytridecan-4-one; D001426:Bacterial Proteins; D007659:Ketones; D012545:Schiff Bases