Isolation of alpha-glucosidase-producing thermophilic bacilli from hot springs of Turkey.

From 42 different hot springs in 6 provinces belonging to distinct geographical regions of Turkey, 451 thermophilic bacilli were isolated and 67 isolates with a high amylase activity were selected to determine the alpha-glucosidase production capacities by using pNPG as a substrate. Alpha-glucosidase production capacities of the isolates varied within the range from 77.18 to 0.001 U/g. Eleven of our thermophilic bacilli produced alpha-glucosidase at significant levels comparable with that of the reference strains tested, thus five strains, F84b (77.18 U/g), A333 (48.64 U/g), F84a (36.64 U/g), E134 (32.09 U/g), and A343 (10.79 U/g) were selected for further experiments. 16S rDNA sequence analysis revealed that these selected isolates all belonged to thermophilic bacilli 16S rDNA genetic group 5, four of them representing the genus Geobacillus, while strain A343 had an uncultured bacterium as the closest relative. Changes of alpha-glucosidase levels in the intracellular and extracellular fractions were determined during 48-h cultivation of A333, A343, F84a, F84b, E134, and the reference strain G. stearothermophilus ATCC 12980. According to alpha-glucosidase production type and enzyme levels in intracellular and extracellular fractions, Geobacillus spp. A333, F84a and F84b were defined as extracellular enzyme producers, whereas the thermophilic bacterium A343 was found to be an intracellular alpha-glucosidase producer, similar to ATCC 12980 strain. Geobacillus sp. E134 differed in alpha-glucosidase production type from all tested isolates and the reference strain; it was described as a membrane-associated cell-bound enzyme producer. In this study, apart from screening a great number of new thermophilic bacilli from the hot springs of Turkey, which have not yet been thoroughly studied, five new thermostable alpha-1,4-glucosidase-producing bacilli that have biotechnological potential with alpha-glucosidases located at different cell positions were obtained.