Partial purification of an esterase produced by B. licheniformis S-86

TORRES SEBASTIÁN; RODRÍGUEZ EMILIO; CASTRO GUILLERMO R.

BIOCELL

IHEM-CONICET

Lugar: Mendoza, Argentina; Año: 2004 vol. 28 p. 59 - 59

0327-9545

Esterases have attracted great interest due to their capacity to synthesize new compounds of high industrial value. A wild-type strain S-86, isolated and characterized in our laboratory, as B. licheniformis is able to produce organic-solvent resistant-esterases. The aim of the present work was the purification of one esterase produced by B. licheniformis S-86. Esterase production was carried out in a 400 ml-airlift reactor type at 50ºC. For enzyme purification the crude extract was precipitate with 50% acetone and then chromatographed in DEAE-cellulose. B. licheniformis S-86 produces two different types esterases; one of them was PMSF-sensitive (type I), but the other esterase activity was not inhibited by PMSF (type II). Precipitation of crude extract with 50% acetone yielded 52%. In the first ion-exchange step, the high esterase activity fractions eluted between 0.20 and 0.40M NaCl with 32% yield. Zymograms revealed that the activity recovered corresponded to type II esterase. In the second ion-exchange step, fractions with esterase activity eluted at 0.20 to 0.25M NaCl. In this last step, type II esterase was purified 6.75-fold with a specific activity of 6.54 U/mg and 10% yield. Concluding, the possibility to obtain pure esterases would allow its application in quiral synthesis of organic compound with potential industrial use. Esterases have attracted great interest due to their capacity to synthesize new compounds of high industrial value. A wild-type strain S-86, isolated and characterized in our laboratory, as B. licheniformis is able to produce organic-solvent resistant-esterases. The aim of the present work was the purification of one esterase produced by B. licheniformis S-86. Esterase production was carried out in a 400 ml-airlift reactor type at 50ºC. For enzyme purification the crude extract was precipitate with 50% acetone and then chromatographed in DEAE-cellulose. B. licheniformis S-86 produces two different types esterases; one of them was PMSF-sensitive (type I), but the other esterase activity was not inhibited by PMSF (type II). Precipitation of crude extract with 50% acetone yielded 52%. In the first ion-exchange step, the high esterase activity fractions eluted between 0.20 and 0.40M NaCl with 32% yield. Zymograms revealed that the activity recovered corresponded to type II esterase. In the second ion-exchange step, fractions with esterase activity eluted at 0.20 to 0.25M NaCl. In this last step, type II esterase was purified 6.75-fold with a specific activity of 6.54 U/mg and 10% yield. Concluding, the possibility to obtain pure esterases would allow its application in quiral synthesis of organic compound with potential industrial use. Esterases have attracted great interest due to their capacity to synthesize new compounds of high industrial value. A wild-type strain S-86, isolated and characterized in our laboratory, as B. licheniformis is able to produce organic-solvent resistant-esterases. The aim of the present work was the purification of one esterase produced by B. licheniformis S-86. Esterase production was carried out in a 400 ml-airlift reactor type at 50ºC. For enzyme purification the crude extract was precipitate with 50% acetone and then chromatographed in DEAE-cellulose. B. licheniformis S-86 produces two different types esterases; one of them was PMSF-sensitive (type I), but the other esterase activity was not inhibited by PMSF (type II). Precipitation of crude extract with 50% acetone yielded 52%. In the first ion-exchange step, the high esterase activity fractions eluted between 0.20 and 0.40M NaCl with 32% yield. Zymograms revealed that the activity recovered corresponded to type II esterase. In the second ion-exchange step, fractions with esterase activity eluted at 0.20 to 0.25M NaCl. In this last step, type II esterase was purified 6.75-fold with a specific activity of 6.54 U/mg and 10% yield. Concluding, the possibility to obtain pure esterases would allow its application in quiral synthesis of organic compound with potential industrial use.