title: Cys139Ser mutation in dimeric nucleoside diphosphate kinase generates catalytically competent monomer.
abstract: Nucleoside diphosphate kinase from a moderate halophile Halomonas sp. 593 (HaNDK) is dimer, while NDK from different origins has been shown to assemble into hexamer, or tetramer. Similar to HaNDK, halophilic NDK from Chromohalobacter salexigens DSM3043 (CsNDK) formed dimeric structure. Cysteine139 conserved between HaNDK and CsNDK is located in the monomer/monomer interface. Substitution of Cys139 for Ser caused dissociation of dimeric CsNDK into monomer in Tris buffer, as determined by field flow fractionation technique. Circular dichroism (CD) profile of the mutant CsNDK was nearly identical to the wild type CsNDK: however, the mutant CsNDK became more susceptible to "endproteinase GluC" cleavage, which could be suppressed by an NDK substrate, ATP. The monomer was enzymatically active, although it is possible that active structure is dimer in the presence of substrate.
authors.affiliation: Applied and Molecular Microbiology, Faculty of Agriculture, Kagoshima University, 1-21-24 Korimoto, Kagoshima 890-0065, Japan.
authors.forename: Hiroko
authors.initials: H
authors.lastname: Tokunaga
authors.affiliation: Alliance Protein Laboratories, 6042 Cornerstone Court West, Suite A, San Diego, CA 92121, USA.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Arakawa
authors.affiliation: Applied and Molecular Microbiology, Faculty of Agriculture, Kagoshima University, 1-21-24 Korimoto, Kagoshima 890-0065, Japan. Electronic address: tokunaga@chem.agri.kagoshima-u.ac.jp.
authors.forename: Masao
authors.initials: M
authors.lastname: Tokunaga
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D002384:Catalysis; D002942:Circular Dichroism; D003545:Cysteine / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D020632:Halomonas / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D009701:Nucleoside-Diphosphate Kinase / Q000235:genetics*
chemical_list: D001426:Bacterial Proteins; D009701:Nucleoside-Diphosphate Kinase; D003545:Cysteine
keywords: ATP; Chromohalobacter salexigens; Dimeric structure; Monomer/monomer interface; Nucleoside diphosphate kinase