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Exquisite Regioselectivity and Increased Transesterification Activity of an Immobilized Bacillus subtilis Protease
Author(s) -
Ferreira Lino,
Ramos M. A.,
Gil Maria Helena,
Dordick Jonathan S.
Publication year - 2002
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp0255457
Subject(s) - chemistry , hydrolysis , protease , bacillus subtilis , immobilized enzyme , subtilisin , biocatalysis , maltose , transesterification , dimethylformamide , lipase , sucrose , organic chemistry , enzyme , chromatography , catalysis , solvent , bacteria , reaction mechanism , biology , genetics
Commercially available proteases and lipases were screened for their ability to acylate regioselectively sucrose with divinyladipate either in pyridine or dimethylformamide (DMF). The protease (EC 3.4.21.62) from Bacillus subtilis (Proleather FG‐F) exhibited the highest conversion (100% in 24 h of reaction in DMF) yielding sucrose 2‐ O ‐vinyladipate as main product. The enzyme preference for a secondary hydroxyl group is a distinct feature of this biocatalyst compared to others described in the literature. Two sets of chemically distinct silica supports were used for Proleather immobilization presenting terminal amino (S APTES ) or hydroxyl groups (S TESPM ‐ pHEMA ). The percentage of immobilized enzyme was smaller in S APTES (7–17%) than in S TESPM ‐ pHEMA (52–56%), yet Proleather immobilized into S APTES supports presented higher total and specific hydrolytic activity. The highest total and specific activities were obtained with S TESPM ‐ pHEMA and S APTES , respectively. Silicas with large pore (bimodal distribution of pores, 130/1200 Å, denoted as S 1000 ) presented higher specific activities relative to those with smaller pore sizes. Furthermore, the synthetic specific activity of S 1000 S APTES immobilized protease was ca. 10‐fold higher than that of the free enzyme. In addition to sucrose, the immobilized protease was used to acylate methyl α‐ d ‐glucopyranoside, trehalose, and maltose in nearly anhydrous DMF. Finally, immobilized Proleather was reasonably stable, retaining ca. 55% activity after six reaction cycles.