Purification and characterization of phosphomannomutase/phosphoglucomutase from Pseudomonas aeruginosa involved in biosynthesis of both alginate and lipopolysaccharide

R. W. Ye, N. A. Zielinski, A. M. Chakrabarty*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

78 Scopus citations

Abstract

The algC gene from Pseudomonas aeruginosa has been shown to encode phosphomannomutase (PMM), an essential enzyme for biosynthesis of alginate and lipopolysaccharide (LPS). This gene was overexpressed under control of the tac promoter, and the enzyme was purified and its substrate specificity and metal ion effects were characterized. The enzyme was determined to be a monomer with a molecular mass of 50 kDa. The enzyme catalyzed the interconversion of mannose 1-phosphate (M1P) and mannose 6-phosphate, as well as that of glucose 1-phosphate (G1P) and glucose 6-phosphate. The apparent K(m) values for M1P and G1P were 17 and 22 μM, respectively. On the basis of K(cat)/K(m) ratio, the catalytic efficiency for G1P was about twofold higher than that for M1P. PMM also catalyzed the conversion of ribose 1-phosphate and 2-deoxyglucose 6-phosphate to their corresponding isomers, although activities were much lower. Purified PMM/phosphoglucomutase (PGM) required Mg2+ for maximum activity; Mn2+ was the only other divalent metal that showed some activation. The presence of other divalent metals in addition to Mg2+ in the reaction inhibited the enzymatic activity. PMM and PGM activities could not be detected in nonmucoid algC mutant strain 8858 and in LPS-rough algC mutant strain AK1012, while they were present in the wild- type strains as well as in algC-complemented mutant strains. This evidence suggests that AlgC functions as PMM and PGM in vivo, converting phosphomannose and phosphoglucose in the biosynthesis of both alginate and LPS.

Original languageEnglish (US)
Pages (from-to)4851-4857
Number of pages7
JournalJournal of bacteriology
Volume176
Issue number16
DOIs
StatePublished - 1994

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

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