Cloning of a mineral phosphate-solubilizing gene from Pseudomonas cepacia

We have recently shown that the ability of some gram-negative bacteria to dissolve poorly soluble calcium phosphates (Mps+ phenotype) is the result of periplasmic oxidation of glucose to gluconic acid via the quinoprotein glucose dehydrogenase (GDH), a component of the direct oxidation pathway. Esch...

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Bibliographic Details
Published in:	Applied and Environmental Microbiology Vol. 61; no. 3; pp. 972 - 978
Main Authors:	Babu-Khan, S. (Beckman Research Institute, Duarte, CA.), Yeo, T.C, Martin, W.L, Duron, M.R, Rogers, R.D, Goldstein, A.H
Format:	Journal Article
Language:	English
Published:	Washington, DC American Society for Microbiology 01-03-1995
Subjects:	Amino Acid Sequence Amino Acid Transport Systems, Basic ATP-Binding Cassette Transporters AZUCARES ACIDOS BACTERIA Bacterial Proteins - biosynthesis Bacterial Proteins - chemistry Bacterial Proteins - genetics Bacteriology Base Sequence Biochemistry Biological and medical sciences Biology of microorganisms of confirmed or potential industrial interest BIOSINTESIS BIOSYNTHESE Biotechnology Burkholderia cepacia - enzymology Burkholderia cepacia - genetics Calcium Phosphates - metabolism CLONACION CLONAGE Cloning, Molecular COMPOSICION QUIMICA COMPOSITION CHIMIQUE Escherichia coli - genetics ESTRUCTURA CELULAR Fundamental and applied biological sciences. Psychology GENE GENES Genes, Bacterial - genetics Genetics Gluconates - metabolism Membrane Proteins Membrane Transport Proteins - genetics MINERALES MINERAUX Mission oriented research Molecular Sequence Data Molecular Weight MUTACION MUTATION Open Reading Frames - genetics Phenotype PROTEINAS PROTEINE PSEUDOMONAS CEPACIA SECUENCIA NUCLEICA Sequence Analysis, DNA Sequence Homology, Amino Acid SEQUENCE NUCLEIQUE SOLUBILIDAD SOLUBILITE Solubility STRUCTURE CELLULAIRE SUCRE ACIDE Gluconic acid Phosphates Calcium Nucleotide sequence Burkholderia cepacia Production Bacteria Oxidation Inorganic element Aminoacid sequence Structural analysis Open reading frame
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Summary:	We have recently shown that the ability of some gram-negative bacteria to dissolve poorly soluble calcium phosphates (Mps+ phenotype) is the result of periplasmic oxidation of glucose to gluconic acid via the quinoprotein glucose dehydrogenase (GDH), a component of the direct oxidation pathway. Escherichia coli K-12 derivatives synthesize apo-GDH but not the cofactor pyrroloquinoline-quinone (PQQ) essential for formation of the holoenzyme. Therefore, in the absence of exogenous PQQ, these strains do not produce gluconic acid and are Mps-. Evidence is presented to show that expression of a single 396-base Pseudomonas cepacia open reading frame (designated gabY) in E. coli JM109 (a K-12 derivative) was sufficient to induce the Mps+ phenotype and production of gluconic acid. We present the nucleotide sequence of this open reading frame which coded for a protein (GabY) with a deduced Mr of 14,235. Coupled transcription-translation of a plasmid (pSLY4 or pGAB1) carrying gabY resulted in production of a protein with an Mr of 14,750. Disruption of the open reading frame of gabY via site-directed mutagenesis changed the phenotype to Mps- and eliminated gluconic acid production. The deduced amino acid sequence of gabY has no apparent homology with those of previously cloned direct oxidation pathway genes but does share regions highly homologous with the histidine permease system membrane-bound protein HisQ as well as other proteins in this family. In the presence of 1 micromolar exogenous PQQ, both JM109(pSLY4) and JM109(pGAB1) produced 10 times as much gluconic acid as was seen with either the plasmid or exogenous PQQ alone. The presence of pGAB1 was also sufficient to cause production of gluconic acid in E. coli HB101 (a K-12-B hybrid). In AG121, an apoGDH-, Tn5 mutant of HB101, the presence of pGAB1 did not cause the production of gluconic acid
Bibliography:	P34 9542735 ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23
ISSN:	0099-2240 1098-5336
DOI:	10.1128/aem.61.3.972-978.1995