Isolation of an Acinetobacter junii SY-01 strain producing an extracellular lipase enantioselectively hydrolyzing Itraconazole precursor, and some properties of the lipase

Abstract

Water-sludge bacteria were screened to find a lipase enantioselectively hydrolyzing itraconazole precursor, which is well known as the starting material of antifungal drug agents. A bacterial strain was isolated and identified as Acinetobacter junii SY-01. After the strain was cultivated, the enzyme was purified 39.4-fold using ultrafiltration and gel filtration through a Sephadex G-100 chromatographic column and the activity yield was 34.9%. The molecular weight of the enzyme was about 40 kDa, as measured by SDS-PAGE, and the optimum pH was 7.0-9.0 and stable at pH 6.0-9.0. The optimum temperature was 45-50degreesC, and 73% of the enzymes activity remained after incubation at 70degreesC for I h. Enzyme activity was enhanced by gall powder, sodium deoxycholate, a cationic detergent Tween 80, and a non-ionic detergent Triton X-100, but was markedly inhibited by metal ions such as Hg2+, Cu2+ Ni2+, Ca2+, and an anionic-surfactant sodium dodecylsulfate. The K. values for (R)-, and (S)-enantiomers of the itraconazole precursor were 0.385 and 21.83 mM, respectively, and the V-max values (muM(.)min(-1)) were 6.73 and 6.49, respectively. The acetyl group among the different acyl moieties of itraconazole precursor showed the highest enantioselectivity for the hydrolysis by the Acinetobacter junii SY-01 lipase, and the lipase from Acinetobacter junii SY-01 displayed better enantioselectivity than that of commercially available lipases and esterases.