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. |
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| Authors: | I S-Correia A Darzins S K Wang A Berry and A M Chakrabarty |
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| Affiliation: | I Sá-Correia, A Darzins, S K Wang, A Berry, and A M Chakrabarty |
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| Abstract: | The specific activities of phosphomannose isomerase (PMI), phosphomannomutase (PMM), GDP-mannose pyrophosphorylase (GMP), and GDP-mannose dehydrogenase (GMD) were compared in a mucoid cystic fibrosis isolate of Pseudomonas aeruginosa and in two spontaneous nonmucoid revertants. In both revertants some or all of the alginate biosynthetic enzymes we examined appeared to be repressed, indicating that the loss of the mucoid phenotype may be a result of decreased formation of sugar-nucleotide precursors. The introduction and overexpression of the cloned P. aeruginosa phosphomannose isomerase (pmi) gene in both mucoid and nonmucoid strains led not only to the appearance of PMI levels in cell extracts several times higher than those present in the wild-type mucoid strain, but also in higher PMM and GMP specific activities. In extracts of both strains, however, the specific activity of GMD did not change as a result of pmi overexpression. In contrast, the introduction of the cloned Escherichia coli manA (pmi) gene in P. aeruginosa caused an increase in only PMI and PMM activities, having no effect on the level of GMP. This suggests that an increase in PMI activity alone does not induce high GMP activity in P. aeruginosa. The heterologous overexpression of the P. aeruginosa pmi gene in the E. coli manA mutant CD1 led to the appearance in cell extracts of not only PMI activity but also GMP activity, both of which are normally undetectable in extracts of CD1. We discuss the implications of these results and propose a mechanism by which overexpression of the P. aeruginosa pmi gene can cause an elevation in both PMM and GMP activities. |
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