title: Expression, crystallization and preliminary X-ray crystallographic analyses of two N-terminal acetyltransferase-related proteins from Thermoplasma acidophilum.
abstract: N-terminal acetylation is one of the most common protein modifications in eukaryotes, occurring in approximately 80-90% of cytosolic mammalian proteins and about 50% of yeast proteins. ARD1 (arrest-defective protein 1), together with NAT1 (N-acetyltransferase protein 1) and possibly NAT5, is responsible for the NatA activity in Saccharomyces cerevisiae. In mammals, ARD1 is involved in cell proliferation, neuronal development and cancer. Interestingly, it has been reported that mouse ARD1 (mARD1(225)) mediates epsilon-acetylation of hypoxia-inducible factor 1alpha (HIF-1alpha) and thereby enhances HIF-1alpha ubiquitination and degradation. Here, the preliminary X-ray crystallographic analyses of two N-terminal acetyltransferase-related proteins encoded by the Ta0058 and Ta1140 genes of Thermoplasma acidophilum are reported. The Ta0058 protein is related to an N-terminal acetyltransferase complex ARD1 subunit, while Ta1140 is a putative N-terminal acetyltransferase-related protein. Ta0058 shows 26% amino-acid sequence identity to both mARD1(225) and human ARD1(235). The sequence identity between Ta0058 and Ta1140 is 28%. Ta0058 and Ta1140 were overexpressed in Escherichia coli fused with an N-terminal purification tag. Ta0058 was crystallized at 297 K using a reservoir solution consisting of 0.1 M sodium acetate pH 4.6, 8%(w/v) polyethylene glycol 4000 and 35%(v/v) glycerol. X-ray diffraction data were collected to 2.17 A. The Ta0058 crystals belong to space group P4(1) (or P4(3)), with unit-cell parameters a = b = 49.334, c = 70.384 A, alpha = beta = gamma = 90 degrees. The asymmetric unit contains a monomer, giving a calculated crystal volume per protein weight (V(M)) of 2.13 A(3) Da(-1) and a solvent content of 42.1%. Ta1140 was also crystallized at 297 K using a reservoir solution consisting of 0.1 M trisodium citrate pH 5.6, 20%(v/v) 2-propanol, 20%(w/v) polyethylene glycol 4000 and 0.2 M sodium chloride. X-ray diffraction data were collected to 2.40 A. The Ta1140 crystals belong to space group R3, with hexagonal unit-cell parameters a = b = 75.174, c = 179.607 A, alpha = beta = 90, gamma = 120 degrees. Two monomers are likely to be present in the asymmetric unit, with a V(M) of 2.51 A(3) Da(-1) and a solvent content of 51.0%.
authors.affiliation: Department of Chemistry, College of Natural Sciences, Seoul National University, Seoul 151-742, South Korea.
authors.forename: Sang Hee
authors.initials: SH
authors.lastname: Han
authors.forename: Jun Yong
authors.initials: JY
authors.lastname: Ha
authors.forename: Kyoung Hoon
authors.initials: KH
authors.lastname: Kim
authors.forename: Sung Jin
authors.initials: SJ
authors.lastname: Oh
authors.forename: Do Jin
authors.initials: DJ
authors.lastname: Kim
authors.forename: Ji Yong
authors.initials: JY
authors.lastname: Kang
authors.forename: Hye Jin
authors.initials: HJ
authors.lastname: Yoon
authors.forename: Se-Hee
authors.initials: SH
authors.lastname: Kim
authors.forename: Ji Hae
authors.initials: JH
authors.lastname: Seo
authors.forename: Kyu-Won
authors.initials: KW
authors.lastname: Kim
authors.forename: Se Won
authors.initials: SW
authors.lastname: Suh
mesh_terms: D000123:Acetyltransferases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000818:Animals; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D001483:Base Sequence; D018360:Crystallography, X-Ray; D017931:DNA Primers; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D063209:N-Terminal Acetyltransferase A; D063213:N-Terminal Acetyltransferase E; D010446:Peptide Fragments / Q000737:chemistry; D013822:Thermoplasma / Q000201:enzymology*
chemical_list: D019843:Archaeal Proteins; D017931:DNA Primers; D010446:Peptide Fragments; D000123:Acetyltransferases; D063209:N-Terminal Acetyltransferase A; C504824:NAA10 protein, human; C508558:Naa10 protein, mouse; D063213:N-Terminal Acetyltransferase E