A new l-glutaminase from Streptomyces pratensis NRC 10: Gene identification, enzyme purification, and characterization.

In the current study, the purified l-glutaminase from Streptomyces pratensis NRC10 (GenBank number KC857622) was characterized. Its molecular weight was estimated to be 46kDa and isoelectric point 7.4. Its Vmax was calculated to be 2.19U/mg/min, while Km was 0.175mM. The optimum pH and temperature were 9 and 45°C, respectively. It was thermostable at 45°C but thermally inactivated at 60°C after 50min. Moreover, its enzymatic activity was enhanced by K+ ions and inhibited by Mg2+ , Cu2+ , Ag+ , Hg2+ , Ni2+ , Fe2+ , Cr2 , Na+ , Ca2+ , and EDTA. A PCR fragment of 1550bp of S. pratensis NRC10 l-glutaminase gene (glsA) was purified and its sequence was determined (GenBank number KJ567136). l-glutaminase from NRC10 was induced mainly by l-glutamic acid. Model 3-D structure was composed of two domains, the serine - dependent beta-lactamase dominant the small STAS domain (Sulphate Transporter and anti-sigma factor antagonist) which had probably functioned as a general NTP binding domain. The two domains are linked by a linker peptide (GLHLMRNPALPGST), but sequence alignment between salt-tolerant glutaminase and the obtained glutaminase showed 44.75% of identity and 57% of similarity. This enzyme appears to have a distinctive structure compared to the rest of glutaminase family, and seems to construct a new subgroup of glutaminase.