Overproduction and assay of Pseudomonas aeruginosa phosphomannose isomerase.

AUTOR(ES)

Gill, J F

RESUMO

Phosphomannose isomerase activity was undetectable in extracts of mucoid (alginate-producing) Pseudomonas aeruginosa. When a P. aeruginosa gene previously shown to complement an alginate-negative mutant was overexpressed under the control of the tac promoter in the broad-host-range controlled-expression vector pMMB22, phosphomannose isomerase activity could be measured in extracts of P. aeruginosa and in a manA (phosphomannose isomerase-negative) mutant of Escherichia coli. P. aeruginosa extracts containing induced levels of enzyme were shown to interconvert fructose 6-phosphate and mannose 6-phosphate. A 56,000-dalton polypeptide was visualized on sodium dodecyl sulfate-polyacrylamide gels after induction in both hosts. When RNA-DNA dot- blot hybridization analysis was used, transcription of algA, the gene coding for P. aeruginosa phosphomannose isomerase, was not measurable from the chromosomes of either mucoid or nonmucoid P. aeruginosa. However, a high level of algA transcription was detected after expression of algA under tac promoter control in pMMB22.

Documentos Relacionados

• Alginate biosynthetic enzymes in mucoid and nonmucoid Pseudomonas aeruginosa: overproduction of phosphomannose isomerase, phosphomannomutase, and GDP-mannose pyrophosphorylase by overexpression of the phosphomannose isomerase (pmi) gene.
• Cloning of Escherichia coli and Pseudomonas aeruginosa phosphomannose isomerase genes and their expression in alginate-negative mutants of Pseudomonas aeruginosa.
• A novel Saccharomyces cerevisiae secretory mutant possesses a thermolabile phosphomannose isomerase.
• Hyphal tip extension in Aspergillus nidulans requires the manA gene, which encodes phosphomannose isomerase.
• Molecular and industrial aspects of glucose isomerase.