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Carboxyl ester hydrolase from P enicillium expansum : cloning, characterization and overproduction by P enicillium griseoroseum
Author(s) -
Corrêa T.L.R.,
Zubieta M.P.,
Teixeira J.A.,
Queiroz M.V.,
Araújo E.F.
Publication year - 2013
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1111/jam.12215
Subject(s) - penicillium expansum , overproduction , penicillium , hydrolase , cloning (programming) , microbiology and biotechnology , biology , fungi imperfecti , biochemistry , enzyme , chemistry , botany , postharvest , computer science , programming language
Aims In this study, a gene that encodes a carboxylesterase ( carb ) in P enicillium expansum GF was cloned, sequenced and overexpressed by P enicillium griseoroseum PG 63, and the enzyme was characterized. Methods and Results The recombinant strain, P . griseoroseum T 55, obtained upon transformation using the plasmid pAN ‐52‐1‐ carb , showed integration of the carb gene into at least two heterologous sites of the genome by S outhern blotting. Furthermore, the recombinant strain T 55 exhibited almost a fourfold increase in carboxylesterase activity compared with PG 63 strain when both were cultured without inducers. Based on the secondary structure and multiple sequence alignments with carboxylesterases, cholinesterase and lipase, a three‐dimensional model was obtained. The α/β barrel topology, that is typical of esterases and lipases, was indicated for the CARB protein with S er 213 ‐ G lu 341 ‐ H is 456 as the putative catalytic triad. CARB preferentially hydrolysed acyl chains with eight carbon atoms, and its activity was optimal at a pH of 7·0 and a temperature of 25°C. CARB exhibited stability in alkaline pH , high activity under mesophilic conditions and stability in organic solvents. Conclusion The CARB protein is potentially useful in bioremediation, food and chemical/pharmaceutical industries. Significance and Impact of the Study This study is the first to report the development of a recombinant strain superproducing a P enicillium sp. carboxylesterase.