title: Characterization of the amino acid residues mediating the unique amino-sugar-1-phosphate acetyltransferase activity of the archaeal ST0452 protein.
abstract: The ST0452 protein from the thermophilic archaean Sulfolobus tokodaii has been identified as an enzyme with multiple sugar-1-phosphate nucleotidylyltransferase and amino-sugar-1-phosphate acetyltransferase (amino-sugar-1-P AcTase) activities. Analysis of the protein showed that in addition to glucosamine-1-phosphate (GlcN-1-P) AcTase activity, it possesses unique galactosamine-1-phosphate (GalN-1-P) AcTase activity not detected in any other proteins. Comparison of the crystal structures of the ST0452 protein and GlmU from Escherichia coli (EcGlmU), which possesses only GlcN-1-P AcTase activity, showed that the overall sequence identity between these two proteins is less than 25 %, but the amino acid residues predicted to comprise the catalytic center of EcGlmU are conserved in the ST0452 protein. To understand the molecular mechanism by which the ST0452 amino-sugar-1-P AcTase activity recognizes two independent substrates, several ST0452 substitution and truncation mutant proteins were constructed and analyzed. We found that His308 is essential for both GalN-1-P and GlcN-1-P AcTase activities, whereas Tyr311 and Asn331 are important only for the GalN-1-P AcTase activity. In addition, deletion of the C-terminal 5 or 11 residues showed that the 11-residue C-terminal region exerts a modest stimulatory effect on GalN-1-P AcTase activity but dramatically suppresses GlcN-1-P AcTase activity. This region also appears to make an important contribution to the thermostability of the entire ST0452 protein. Systematic deletions from the C-terminus also demonstrated that the C-terminal region with the β-helix structure has an important role mediating the trimerization of the ST0452 protein. This is the first report of an analysis of a thermostable archaeal enzyme exhibiting multiple amino-sugar-1-P AcTase activities.
authors.affiliation: State Key Laboratory of Marine Environmental Science, Institute of Marine Microbes and Ecospheres, Xiamen University, Xiamen, 361005, People's Republic of China.
authors.forename: Zilian
authors.initials: Z
authors.lastname: Zhang
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Shimizu
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Kawarabayasi
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry; D005688:Galactosamine / Q000031:analogs & derivatives* / Q000378:metabolism; D005695:Galactosephosphates / Q000378:metabolism*; D051081:Glucosamine 6-Phosphate N-Acetyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D016944:Sulfolobus / Q000201:enzymology*
chemical_list: D019843:Archaeal Proteins; D029968:Escherichia coli Proteins; D005695:Galactosephosphates; C520333:GlmU protein, E coli; D009097:Multienzyme Complexes; C079528:galactosamine 1-phosphate; D005688:Galactosamine; D051081:Glucosamine 6-Phosphate N-Acetyltransferase