Characterization of intracellular β-galactosidase from Bacillussubtilis 4NK and Bacillus paralicheniformis 5NK isolated from ahot water spring and effects of various inhibitors on enzymeactivity

Abstract

In this study, the intracellular β-galactosidases of Bacillus subtilis 4NK and Bacillusparalicheniformis 5NK isolated from Bingöl Binkap hot spring was partially purified andcharacterized. As a result of purification, the yield of the enzyme for B. subtilis 4NK was85.2% and the purification fold was 2.8. The yield for B. paralicheniformis 5NK was76.8% and the purification fold was 2.0. The optimum temperature of the enzyme wasdetermined as 45 oC for B. subtilis 4NK and 55 oC for B. paralicheniformis 5NK and theoptimum pH was 6.0 for both. In addition, in the thermal stability experiments even atthe end of 120 min both enzymes were stable at 50 oC. It was determined that thepartially purified enzyme activity increased in the presence of iodoacetamide andphenylmethylsulfonylfluoride for B. subtilis 4NK, dithiothreitol, N-ethylenemaleimide and phenylmethylsulfonylfluoride for B. paralicheniformis 5NK. The metals were foundto activate the enzyme at low concentrations of Co2+, Cd2+ and Mn2+ for B. subtilis 4NK,Cu2+ and Cd2+ were found to inhibit the enzyme at high rates for B. paralicheniformis 5NK. Km and Vmax values for 4NK and 5NK, respectively; 23.80 mM, 1.978 µmol/min and 5.61 mM, 1.869 µmol/min.