~One-hundred-and-f ifty-two isolates of the genus Thermus, collected from hot springs on four continents, were screened for evidence of the presence of the thermophilic Type II restriction endonuclease Taql (TKGA). The presence of isoschizomers of Taql in 27 of the isolates, originating from hot springs in New Zealand, Iceland, USA, Japan, mainland Portugal and the island of S%o Miguel in the Azores, is reported. Six of the Taql-containing isolates from diverse geographical locations, identified by means of DNNDNA homology and 165 rRNA sequence alignment as belonging to the Thermus species T. aquaticus, T. filiformis, T. thermophilus, T. scotoductus and T. brockianus, were selected for comparative studies. The Taql isoschizomers from each of the six isolates were partially purified. They differed in their magnesium ion requirements, isoelectric points, subunit molecular masses and thermal stability.
Current study was initiated to investigate steroid hydroxylation potential of a new food born thermophile Geobacillus kaustophilus. Incubation of progesterone and testosterone with G. kaustophilus resulted in stereospecific hydroxylation of progesterone and testosterone giving monohydroxylated and dihydroxylated products. A cleavage of ring B of the steroid nucleus was also obtained which is a novel metabolite. Geobacillus kaustophilus has not been used before for steroid hydroxylations. Thermophilic enzymes have a significant advantage for their isolation at elevated temperatures and can be used for cell free transformations offering a convenient method for introduction of hydroxyl groups in a stereospecific and regiospecific manner into steroid nuclei. Stereochemical oxygenation of a steroid nucleus is not possible with chemical reactions and microbial hydroxylation of steroids may have a direct impact on drug development.ConclusionG. kaustophilus has an excellent potential for stereospecific hydroxylation of natural steroid nucleus at elevated temperature. This study was supported by Research Administration, Kuwait University, Kuwait.
Isolates of thermophilic bacteria from desert soil in Kuwait, heavily contaminated with crude oil, have been screened for the presence of restriction endonuclease activity. One of the isolates (B7S), identified as Bacillus stearothermophilus, showed a high level of restriction endonuclease activity when a cell-free extract was incubated with lambda bacteriophage DNA at 65 degrees C. A type II restriction endonuclease (BstB7SI) has been partially purified from this isolate. BstB7SI recognises the six-base sequence RCCGGY (R = A or G; Y = T or C) and hydrolyses the phosphodiester bond in both strands of the DNA substrate between the first and second bases of the recognition sequence 5'-R decreases CCGGY-3'producing four-base 5' overhangs. BstB7SI is therefore an isoschizomer of the mesophilic prototype restriction endonuclease Cfr10I. BstB7SI has a pH optimum of 9.7, requires 10 mM MgCl2 and 75 mM NaCl for maximum activity, and retains full enzyme activity when incubated for 5 min at temperatures up to 70 degrees C.
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