Structure of the adenylation domain Thr1 involved in the biosynthesis of 4‐chlorothreonine in Streptomyces sp. OH‐5093—protein flexibility and molecular bases of substrate specificity. (7th August 2017)

• Record Type: Journal Article
• Authors: Scaglione, Antonella; Fullone, Maria Rosaria; Montemiglio, Linda Celeste; Parisi, Giacomo; Zamparelli, Carlotta; Vallone, Beatrice; Savino, Carmelinda; Grgurina, Ingeborg
• Is Part Of: FEBS journal. Volume 284:Number 18(2017)
• Page Start: 2981
• Page End: 2999
• Publication Date: 2017-08-07
• Subjects: adenylation domain -- crystallography -- kinetic analysis -- nonribosomal code -- substrate specificity; Biochemistry -- Periodicals; Molecular biology -- Periodicals; Pathology, Molecular -- Periodicals; 572
• DOI: 10.1111/febs.14163
• Languages: English
• ISSNs: 1742-464X

Abstract

Abstract : We determined the crystal structure of Thr1, the self‐standing adenylation domain involved in the nonribosomal‐like biosynthesis of free 4‐chlorothreonine in Streptomyces sp. OH‐5093. Thr1 shows two monomers in the crystallographic asymmetric unit with different relative orientations of the C‐ and N‐terminal subdomains both in the presence of substrates and in the unliganded form. Cocrystallization with substrates, adenosine 5′‐triphosphate andl ‐threonine, yielded one monomer containing the two substrates and the other in complex withl ‐threonine adenylate, locked in a postadenylation state. Steady‐state kinetics showed that Thr1 activatesl ‐Thr and its stereoisomers, as well asd ‐Ala, l ‐ andd ‐Ser, albeit with lower efficiency. Modeling of these substrates in the active site highlighted the molecular bases of substrate discrimination. This work provides the first crystal structure of a threonine‐activating adenylation enzyme, a contribution to the studies on conformational rearrangement in adenylation domains and on substrate recognition in nonribosomal biosynthesis. Database: Structural data are available in the Protein Data Bank under the accession number5N9W and5N9X . Abstract : The first crystal structure of a threonine‐activating adenylation domain, Thr1, was obtained in the ligand‐free and ligand‐bound form, both containing two independent chains in the asymmetric unit. The structure arising from cocrystallization with substrates shows one conformation bound to adenosine 5′‐triphosphate andl ‐threonine, the other tol ‐Thr‐adenylate. The analysis of substrate specificity provided insight into the structural determinants of substrate recognition.