title: Structural characterization of a mitochondrial 3-ketoacyl-CoA (T1)-like thiolase from Mycobacterium smegmatis.
abstract: Thiolases catalyze the degradation and synthesis of 3-ketoacyl-CoA molecules. Here, the crystal structures of a T1-like thiolase (MSM-13 thiolase) from Mycobacterium smegmatis in apo and liganded forms are described. Systematic comparisons of six crystallographically independent unliganded MSM-13 thiolase tetramers (dimers of tight dimers) from three different crystal forms revealed that the two tight dimers are connected to a rigid tetramerization domain via flexible hinge regions, generating an asymmetric tetramer. In the liganded structure, CoA is bound to those subunits that are rotated towards the tip of the tetramerization loop of the opposing dimer, suggesting that this loop is important for substrate binding. The hinge regions responsible for this rotation occur near Val123 and Arg149. The Lα1-covering loop-Lα2 region, together with the Nβ2-Nα2 loop of the adjacent subunit, defines a specificity pocket that is larger and more polar than those of other tetrameric thiolases, suggesting that MSM-13 thiolase has a distinct substrate specificity. Consistent with this finding, only residual activity was detected with acetoacetyl-CoA as the substrate in the degradative direction. No activity was observed with acetyl-CoA in the synthetic direction. Structural comparisons with other well characterized thiolases suggest that MSM-13 thiolase is probably a degradative thiolase that is specific for 3-ketoacyl-CoA molecules with polar, bulky acyl chains.
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore, Karnataka 560 012, India.
authors.forename: Neelanjana
authors.initials: N
authors.lastname: Janardan
authors.affiliation: Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, University of Oulu, FIN-90014 Oulu, Finland.
authors.forename: Rajesh K
authors.initials: RK
authors.lastname: Harijan
authors.affiliation: Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, University of Oulu, FIN-90014 Oulu, Finland.
authors.forename: Tiila Riikka
authors.initials: TR
authors.lastname: Kiema
authors.affiliation: Faculty of Biochemistry and Molecular Medicine, Biocenter Oulu, University of Oulu, FIN-90014 Oulu, Finland.
authors.forename: Rikkert K
authors.initials: RK
authors.lastname: Wierenga
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore, Karnataka 560 012, India.
authors.forename: M R N
authors.initials: MR
authors.lastname: Murthy
mesh_terms: D000102:Acetyl-CoA C-Acyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D007700:Kinetics; D008928:Mitochondria / Q000737:chemistry* / Q000201:enzymology; D024101:Mitochondrial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020102:Mycobacterium smegmatis / Q000737:chemistry* / Q000145:classification / Q000201:enzymology; D010802:Phylogeny; D011485:Protein Binding; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D024101:Mitochondrial Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D000102:Acetyl-CoA C-Acyltransferase
keywords: Mycobacterium smegmatis; coenzyme A; crystal structure; thiolase