title: Promiscuity of Exiguobacterium sp. AT1b o-succinylbenzoate synthase illustrates evolutionary transitions in the OSBS family.
abstract: Catalytic promiscuity, which is the ability to catalyze more than one reaction in the same active site, is thought to facilitate the evolution of new protein functions. Although many enzymes are catalytically promiscuous, there is little direct evidence to show how promiscuous activities evolved into biological functions. We are seeking evidence for this model by studying the o-succinylbenzoate synthase (OSBS) family. Most enzymes within this family only catalyze OSBS, which is a step in menaquinone synthesis. However, several characterized enzymes in one branch of the family (called the NSAR/OSBS subfamily) efficiently catalyze both OSBS and N-succinylamino acid racemization (NSAR). Based on genome context, NSAR appears to be the only biological function of some characterized NSAR/OSBS enzymes, while both activities are biologically relevant in others. The promiscuity model predicts that these enzymes evolved from an ancestral OSBS which promiscuously catalyzed NSAR as a side reaction that was not biologically relevant. If so, the model predicts that some extant OSBS enzymes would have low levels of promiscuous NSAR activity. This manuscript describes such an enzyme from Exiguobacterium sp. AT1b (ExiOSBS). We show that ExiOSBS efficiently catalyzes OSBS (kcat/KM=2.6×10(6) M(-1) s(-1)), but its efficiency for the NSAR reaction is only 41 M(-1) s(-1). Moreover, genome context indicates that OSBS is the only biologically relevant activity. ExiOSBS diverged from the NSAR/OSBS subfamily before NSAR emerged as a biologically relevant activity. These results provide evidence that NSAR activity originated as a promiscuous activity in an ancestor of the NSAR/OSBS subfamily.
authors.affiliation: Department of Biochemistry and Biophysics, Texas A&M University, 2128 TAMU, College Station, TX 77843-2128, United States. Electronic address: ashleybrizendine@neo.tamu.edu.
authors.forename: Ashley M
authors.initials: AM
authors.lastname: Brizendine
authors.affiliation: Department of Biochemistry and Biophysics, Texas A&M University, 2128 TAMU, College Station, TX 77843-2128, United States. Electronic address: denis.odokos@gmail.com.
authors.forename: Denis
authors.initials: D
authors.lastname: Odokonyero
authors.affiliation: Department of Biochemistry and Biophysics, Texas A&M University, 2128 TAMU, College Station, TX 77843-2128, United States. Electronic address: amcmillan@tamu.edu.
authors.forename: Andrew W
authors.initials: AW
authors.lastname: McMillan
authors.affiliation: Natural Products LINCHPIN Laboratory, Texas A&M University, 3255 TAMU, College Station, TX 77840, United States. Electronic address: mingzhao.zhu@mail.chem.tamu.edu.
authors.forename: Mingzhao
authors.initials: M
authors.lastname: Zhu
authors.affiliation: Natural Products LINCHPIN Laboratory, Texas A&M University, 3255 TAMU, College Station, TX 77840, United States. Electronic address: ken.hull@chem.tamu.edu.
authors.forename: Ken
authors.initials: K
authors.lastname: Hull
authors.affiliation: Natural Products LINCHPIN Laboratory, Texas A&M University, 3255 TAMU, College Station, TX 77840, United States; Department of Chemistry, Texas A&M University, 3255 TAMU, College Station, TX 77840, United States. Electronic address: romo@chem.tamu.edu.
authors.forename: Daniel
authors.initials: D
authors.lastname: Romo
authors.affiliation: Department of Biochemistry and Biophysics, Texas A&M University, 2128 TAMU, College Station, TX 77843-2128, United States. Electronic address: margy.glasner@tamu.edu.
authors.forename: Margaret E
authors.initials: ME
authors.lastname: Glasner
mesh_terms: D056449:Bacillales / Q000201:enzymology* / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites; D019755:Carbon-Carbon Lyases / Q000737:chemistry* / Q000235:genetics*; D002384:Catalysis; D004789:Enzyme Activation; D019143:Evolution, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D013379:Substrate Specificity
chemical_list: D019755:Carbon-Carbon Lyases; C052986:o-succinylbenzoic acid synthase
keywords: Catalytic promiscuity; Enzymes; Protein evolution