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

Abstract

In the current study, the purified L-glutaminase from Streptomyces pratensis NRC10 (GenBank number KC857622) was characterized. Its molecular weight was estimated to be 46 kDa and isoelectric point 7.4. Its Vmax was calculated to be 2.19 U/mg/min, while Km was 0.175 mM. The optimum pH and temperature were 9 and 45 °C, respectively. It was thermostable at 45 °C but thermally inactivated at 60 °C after 50 min. 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 1550 bp 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.