Phytase Activity In Certain Groups Of Bacteria. Search For And Cloning Of Genes Of Bacterial Phytases

摘要

A search was made for phytase genes in nine strains of Bacillus from the collection of the Institute of Molecular Genetics of the Russian Academy of Sciences. The optimum growth of the strains IX-22, IX-12B, K17-2, K18, IMG I, IMG II, M4, and M8 occurs at 50-60 deg C, and for strain 790 at 45-47 deg C. According to the data of an analysis of the 16S RNA gene, Bacillus sp. 790 belongs to the B. subtilislamyloliquefaciens group. The remaining eight strains were identified as B. licheniformis. The search for phytase genes in the enumerated strains was performed by PCR using primers constructed on the basis of the conservative sequences of the phyA gene of B. amyloliquefaciens FZB45; bacillar chromosomal DNAs were used as the templates. The nucleotide sequences of the PCR fragments obtained proved to be highly homologous to the known genes of bacillar phytases. Gene libraries of B. licheniformis M8 and B. amyloliquefaciens 790 were constructed in E. coli; clones containing phytase genes were selected from them. A total of 24 strains of Pseudomonas of various species, five strains of Xanthomonas maltophilia, and one strain of Xanthomonas malvacearum from the same collection were tested for phytase activity, which was determined in thirteen strains of Pseudomonas and six strains of Xanthomonas. It was shown that in Pseudomonas the accumulation of phytase occurs at late stages of growth (more than 2 days). The optimum pH was determined for phytases from three strains of Pseudomonas. It was found that the enzymes are the most active at pH 5.5.