Mineral phosphate solubilization by rhizosphere bacteria and scope for manipulation of the direct oxidation pathway involving glucose dehydrogenase.

Microbial biodiversity in the soil plays a significant role in metabolism of complex molecules, helps in plant nutrition and offers countless new genes, biochemical pathways, antibiotics and other metabolites, useful molecules for agronomic productivity. Phosphorus being the second most important macro-nutrient required by the plants, next to nitrogen, its availability in soluble form in the soils is of great importance in agriculture. Microbes present in the soil employ different strategies to make use of unavailable forms of phosphate and in turn also help plants making phosphate available for plant use. Azotobacter, a free-living nitrogen fixer, is known to increase the fertility of the soil and in turn the productivity of different crops. The glucose dehydrogenase gene, the first enzyme in the direct oxidation pathway, contributes significantly to mineral phosphate solubilization ability in several Gram-negative bacteria. It is possible to enhance further the biofertilizer potential of plant growth-promoting rhizobacteria by introducing the genes involved mineral phosphate solubilization without affecting their ability to fix nitrogen or produce phytohormones for dual benefit to agricultural crops. Glucose dehydrogenases from Gram-negative bacteria can be engineered to improve their ability to use different substrates, function at higher temperatures and EDTA tolerance, etc., through site-directed mutagenesis.

Glucose dehydrogenase of a rhizobacterial strain of Enterobacter asburiae involved in mineral phosphate solubilization shares properties and sequence homology with other members of enterobacteriaceae.

Glucose dehydrogenase (GDH) of Gram-negative bacteria is a membrane bound enzyme catalyzing the oxidation of glucose to gluconic acid and is involved in the solubilization of insoluble mineral phosphate complexes. A 2.4 kb glucose dehydrogenase gene (gcd) of Enterobacter asburiae sharing extensive homology to the gcd of other enterobacteriaceae members was cloned in a PCR-based directional genome walking approach and the expression confirmed in Escherichia coli YU423 on both MacConkey glucose agar and hydroxyapatite (HAP) containing media. Mineral phosphate solubilization by the cloned E. asburiae gcd was confirmed by the release of significant amount of phosphate in HAP containing liquid medium. gcd was over expressed in E. coli AT15 (gcd::cm) and the purified recombinant protein had a high affinity to glucose, and oxidized galactose and maltose with lower affinities.The enzyme was highly sensitive to heat and EDTA, and belonged to Type I, similar to GDH of E. coli.