Alginic acid synthesis in Pseudomonas aeruginosa mutants defective in carbohydrate metabolism

Abstract

Mutant cells of mucoid Pseudomonas aeruginosa isolated from cystic fibrosis patients were examined for their ability to synthesize alginic acid in resting cell suspensions. Unlike the wild-type strain which synthesizes alginic acid from glycerol, fructose, mannitol, glucose, gluconate, glutamate, or succinate, mutants lacking specific enzymes of carbohydrate metabolism are uniquely impaired. A phosphoglucose isomerase mutant did not synthesize the polysaccharide from mannitol, nor did a glucose 6-phosphate dehydrogenase mutant synthesize the polysaccharide from mannitol or glucose. Mutants lacking the Entner-Doudoroff pathway dehydrase or aldolase failed to produce alginate from mannitol, glucose, or gluconate, as a 3-phosphoglycerate kinase or glyceraldehyde 3-phosphate dehydrogenase mutant failed to produce from glutamate or succinate. These results demonstrate the primary role of the Entner-Doudoroff pathway enzymes in the synthesis of alginate from glucose, mannitol, or gluconate and the role of glyceraldehyde 3-phosphate dehydrogenase reaction for the synthesis from gluconeogenic precursors such as glutamate. The virtual absence of any activity of phosphomannose isomerase in cell extracts of several independent mucoid bacteria and the impairment of alginate synthesis from mannitol in mutants lacking phosphoglucose isomerase or glucose 6-phosphate dehydrogenase rule out free mannose 6-phosphate as an intermediate in alginate biosynthesis. grown overnight in the synthetic medium described by 238 on August 4, 2020 by guest

Influence of nutrient media on the chemical composition of the exopolysaccharide from mucoid and non-mucoid Pseudomonas aeruginosa

“…The differences noted in the chemical composition of the alginate produced when the nutrient components were varied in the culture media may be due to either regulation of the genes controlling biosynthesis of alginate or the lack of enzymes involved in carbohydrate metabolism. This latter mechanism has been demonstrated by Banerjee [35,36] and could explain our results obtained with the two strains isolated from bronchiectasis when grown in five complex media. However, there are several widely dispersed genes involved in regulation and synthesis of algihate that are influenced by environmental factors [14,15,17] such as osmotic stress [13], ethanol dehydratation [2], oxygen tensions [37] and nitrogen starvation [16].…”

The Entner-Doudoroff pathway: history, physiology and molecular biology

“…Alginate, a polysaccharide derived from the metabolic precursor fructose-6-phosphate, is formed during growth on glycerol, fructose, mannitol, glucose, gluconate, or succinate. Phosphoglucose isomerase, glucose-6-phosphate dehydrogenase, EDD-, and EDA-mutants are all defective in alginate synthesis [58], as are fructose 1,6-bisphosphate aldolase mutants [59]. Labelling studies indicate that the majority of labelled carbon in alginate comes not from the pyruvate formed during glucose catabolism, but from glyceraldehyde-3-phosphate only [60].…”

Mode of action of recombinant Azotobacter vinelandii mannuronan C‐5 epimerases AlgE2 and AlgE4

“…The mannuronan will be labeled both in the 5 and 2 positions since the 3 H‐5 glucose is partly metabolized through the Entner–Duodoroff pathway 28. The relative distribution of tritium label in these two positions in the sample was determined by epimerization with AlgE4, and the reaction was monitored both by tritium release and by NMR spectroscopy.…”