title: The phosphofructokinase-B (MJ0406) from Methanocaldococcus jannaschii represents a nucleoside kinase with a broad substrate specificity.
abstract: Recently, unusual non-regulated ATP-dependent 6-phosphofructokinases (PFK) that belong to the PFK-B family have been described for the hyperthermophilic archaea Desulfurococcus amylolyticus and Aeropyrum pernix. Putative homologues were found in genomes of several archaea including the hyperthermophilic archaeon Methanocaldococcus jannaschii. In this organism, open reading frame MJ0406 had been annotated as a PFK-B sugar kinase. The gene encoding MJ0406 was cloned and functionally expressed in Escherichia coli. The purified recombinant enzyme is a homodimer with an apparent molecular mass of 68 kDa composed of 34 kDa subunits. With a temperature optimum of 85 degrees C and a melting temperature of 90 degrees C, the M. jannaschii nucleotide kinase represents one of the most thermoactive and thermostable members of the PFK-B family described so far. The recombinant enzyme was characterized as a functional nucleoside kinase rather than a 6-PFK. Inosine, guanosine, and cytidine were the most effective phosphoryl acceptors. Besides, adenosine, thymidine, uridin and xanthosine were less efficient. Extremely low activity was found with fructose-6-phosphate. Further, the substrate specificity of closely related PFK-Bs from D. amylolyticus and A. pernix were reanalysed.
authors.affiliation: Institut für Allgemeine Mikrobiologie, Christian-Albrechts-Universität Kiel, Am Botanischen Garten 1-9, 24118, Kiel, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Hansen
authors.forename: Linda
authors.initials: L
authors.lastname: Arnfors
authors.forename: Rudolf
authors.initials: R
authors.lastname: Ladenstein
authors.forename: Peter
authors.initials: P
authors.lastname: Schönheit
mesh_terms: D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D019641:DNA, Archaeal; D004795:Enzyme Stability; D007700:Kinetics; D017017:Methanococcus / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D025221:Phosphofructokinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010770:Phosphotransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D019843:Archaeal Proteins; D019641:DNA, Archaeal; D021122:Protein Subunits; D011994:Recombinant Proteins; D010770:Phosphotransferases; D025221:Phosphofructokinases; C021892:nucleoside phosphotransferase