5wsy

The complex structure of SAV606 with N-carboxymethyl-3-aminobutyrate

spinqualitylabelsall models 5wsy, resolution 2.00Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 5wsy is a 2 chain structure with sequence from Streptomyces avermitilis MA-4680 = NBRC 14893. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Method: X-ray diffraction, Resolution 2Å Ligands: 7UC Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Function Q79ZR9_STRAW Publication Abstract from PubMed Thioesterases catalyze hydrolysis of acyl thioesters to release carboxylic acid or macrocyclization to produce the corresponding macrocycle in the biosynthesis of fatty acids, polyketides, or nonribosomal peptides. Recently, we reported that the thioesterase CmiS1 from Streptomyces sp. MJ635-86F5 catalyzes the Michael addition of glycine to an alpha,beta-unsaturated fatty acyl thioester followed by thioester hydrolysis in the biosynthesis of the macrolactam antibiotic cremimycin. However, the molecular mechanisms of CmiS1-catalyzed reactions are unclear. Here, we report on the functional and structural characterization of the CmiS1 homolog SAV606 from Streptomyces avermitilis MA-4680. In vitro analysis indicated that SAV606 catalyzes the Michael addition of glycine to crotonic acid thioester and subsequent hydrolysis yielding (R)-N-carboxymethyl-3-aminobutyric acid. We also determined the crystal structures of SAV606 both in ligand-free form at 2.4 A resolution and in complex with (R)-N-carboxymethyl-3-aminobutyric acid at 2.0 A resolution. We found that SAV606 adopts an alpha/beta hotdog fold and has an active site at the dimeric interface. Examining the complexed structure, we noted that the substrate-binding loop comprising Tyr-53-Asn-61 recognizes the glycine moiety of (R)-N-carboxymethyl-3-aminobutyric acid. Moreover, we found that SAV606 does not contain an acidic residue at the active site, which is distinct from canonical hotdog thioesterases. Site-directed mutagenesis experiments revealed that His-59 plays a crucial role in both the Michael addition and hydrolysis via a water molecule. These results allow us to propose the reaction mechanism of the SAV606-catalyzed Michael addition and thioester hydrolysis and provide new insight into the multiple functions of a thioesterase family enzyme. Structural analysis of the dual-function thioesterase SAV606 unravels the mechanism of Michael addition of glycine to an alpha,beta-unsaturated thioester.,Chisuga T, Miyanaga A, Kudo F, Eguchi T J Biol Chem. 2017 Jun 30;292(26):10926-10937. doi: 10.1074/jbc.M117.792549. Epub , 2017 May 18. PMID:28522606 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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