Bacillus subtilis ORF yybQ encodes a manganese-dependent inorganic pyrophosphatase with distinctive properties: the first of a new class of soluble pyrophosphatase?
The N-terminal 15 amino acids of the major protein associated with inorganic pyrophosphatase activity in Bacillus subtilis WB600 are identical to those of B. subtilis ORF yybQ. This ORF was amplified from B. subtilis WB600 DNA by PCR and cloned into an overexpression vector in Escherichia coli. Induction of overexpression produced a soluble protein of 34000 Da by SDS-PAGE and by matrix-assisted laser desorption and ionization mass spectrometry. The overexpressed protein had a high specific activity for the hydrolysis of magnesium pyrophosphate, and was specifically and reversibly activated by Mn2+ ions. These properties are identical to those of inorganic pyrophosphatase purified from B. subtilis WB600. No significant similarity was found between the derived sequence of the B. subtilis yybQ-encoded protein and published sequences of identified inorganic pyrophosphatases of Eukarya, Bacteria or Archaea domains. However, there is significant similarity to three putative proteins of unknown function from the archaea Methanococcus jannaschii and Archaeoglobus fulgidus, and from Streptococcus gordonii. The genomes of B. subtilis, M. jannaschii and A. fulgidus do not contain sequences similar to those of hitherto known soluble inorganic pyrophosphatases. The present findings, together with a survey of the properties of inorganic pyrophosphatases from 38 different sources, suggest that the B. subtilis yybQ-encoded protein is the first fully characterized member of a new class of inorganic pyrophosphatase.
van AlebeekG.-J.,
KeitjensJ.T.,
vander DriftC.1994; Purification and characterization of inorganic pyrophosphatase from Methanobacterium thermoautotrophicum.
. Biochim Biophys Acta 1206:231–239
BaykovA.A.,
ArtjukovA.A.,
AvaevaS.1976; Fluoride inhibition of inorganic pyrophosphatase. I. Kinetic studies in a Mg2+-PPi system using a new continuous enzyme assay.. Biochim Biophys Acta 429:982–992
BaykovA.A.,
BakulevaN.P.,
NazarovaT.I.,
AvaevaS.M.1977; Fluoride inhibition of inorganic pyrophosphatase. II. Isolation and characterization of a covalent intermediate between enzyme and entire substrate molecule.. Biochim Biophys Acta 481:184–194
BaykovA.A.,
AlexandrovA.P.,
SmirnovaI.N.1992; A two- step mechanism of fluoride inhibition of rat liver inorganic pyrophosphatase.. Arch Biochem Biophys 294:238–243
BradfordM.M.1976; A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.. Anal Biochem 72:248–254
CoopermanB.S.,
AlexanderA.B.,
LahtiR.1992; Evolutionary conservation of the active site of soluble inorganic pyrophosphatase.. Trends Biochem Sci 17:262–266
FleischmannR.D.,
AdamsM.D.,
WhiteO.
37 other authors 1995; Whole-genome random sequencing and assembly of Haemophilus influenzae Rd.. Science 269:496–512
HachimoriA.,
TakedaA.,
KaibuchiM.,
OhkawaraN.Samejima,
SamejimaT.1975; Purification and characterization of inorganic pyrophosphatase from B. stearothermophilus.
. J Biochem 77:1177–1183
HimmelreichR.,
HilbertH.,
PlagensH.,
PirkIE.,
LiB.-C.,
HerrmannR.1996; Complete sequence analysis of the genome of the bacterium Mycoplasma pneumoniae.
. Nucleic Acids Res 24:4420–4449
du JardinP.,
Rojas-BeltranJ.,
GebhardtC.,
BrasseurR.1995; Molecular cloning and characterization of a soluble inorganic pyrophosphatase in potato.. Plant Physiol 109:853–860
KolakowskiL.F.,
SchloesserM.,
CoopermanB.S.
1988; Cloning, molecular characterization and chromosome localization of the inorganic pyrophosphatase (PPA) gene from Saccharomyces cerevisiae.
. Nucleic Acids Res 16:10441–10452
KuhnN.J.,
WardS.1998; Purification, properties, and multiple forms of a manganese-activated inorganic pyrophosphatase from Bacillus subtilis.
. Arch Biochem Biophys 354:47–56
KunstF.,
OgasawaraN.,
MoszerI.
148 other authors 1997; The complete genome sequence of the Gram-positive bacterium Bacillus subtilis.
. Nature 390:249–256
KuranovaI.P.,
ObmolovaG.V.,
KonarevaN.V.
1987; Inorganic pyrophosphatase from the extremely thermophilic bacterium Thermus thermophilus HB8.. Dokl Akad Nauk SSSR 295:1013–1016
MitchellS.J.,
MinnickM.F.1997; Cloning, functional expression, and complementation analysis of an inorganic pyrophosphatase from Bartonella bacilliformis.
. Can J Microbiol 43:734–743
OgasawaraN.,
NakaiS.,
YoshikawaH.1994; Systematic sequencing of the 180 kilobase region of the B. subtilis chromosome containing the replication origin.. DNA Res 1:1–14
RichterO.-M.,
ScháferG.1992a; . Purification and enzymic characterization of the cytoplasmic pyrophosphatase from the thermoacidophilic archaebacterium Thermoplasma acidophilum.
. Eur J Biochem 209:343–349
RichterO.M.,
ScháferG.1992b; . Cloning and sequencing of the gene for the cytoplasmic inorganic pyrophosphatase from the thermoacidophilic archaebacterium Thermoplasma acidophilum. Eur
. J Biochem 209:351–355
StarkM.J.R.,
MilnerJ.S.
1989; Cloning and analysis of the Kluyueromyces lactis TRPl gene: a chromosomal locus flanked by genes encoding inorganic pyrophosphatase and histone H3.. Yeast 5:35–50
ThompsonJ.D.,
HigginsD.G.,
GibsonT.J.
1994; CLUSTAL w: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice.. Nucleic Acids Res 22:4673–680
TonoH.,
KornbergA.1967a; Biochemical studies of bacterial sporulation. IV. Inorganic pyrophosphatase of vegetative cells and spores of Bacillus megaterium.
. J Bacteriol 93:1819–1824
TonoH.,
KornbergA.1967b; Biochemical studies of bacterial sporulation III. Inorganic pyrophosphatase of vegetative cells and spores of Bacillus subtilis.
. J Biol Chem 242:2375–2382
VolkS.E.Baykov,
BaykovA.1984; Isolation and subunit composition of membrane inorganic pyrophosphatase from rat liver mitochondria.. Biochim Biophys Acta 791:198–204
WhittakerC.J.,
ClemensD.L.,
KolenbranderD.L.
1996; Insertional inactivation of an intrageneric coaggregation-relevant adhesin locus from Streptococcus gordonii DLl (Challis).. Infect Immun 4:4137–4142
WongS.C.K.,
HallD.C.,
JosseJ.1970; Constitutive inorganic pyrophosphatase of Escherichia coli. III. Molecular weight and physical properties of the enzyme and its subunits.. J Biol Chem 245:4335–4345
Bacillus subtilis ORF yybQ encodes a manganese-dependent inorganic pyrophosphatase with distinctive properties: the first of a new class of soluble pyrophosphatase?