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Biocatalysis with E scherichia coli ‐overexpressing cyclopentanone monooxygenase immobilized in polyvinyl alcohol gel
Author(s) -
Rebroš M.,
Lipták L.,
Rosenberg M.,
Bučko M.,
Gemeiner P.
Publication year - 2014
Publication title -
letters in applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.698
H-Index - 110
eISSN - 1472-765X
pISSN - 0266-8254
DOI - 10.1111/lam.12227
Subject(s) - biocatalysis , monooxygenase , polyvinyl alcohol , escherichia coli , bioconversion , cyclopentanone , chemistry , recombinant dna , biotransformation , enzyme , biochemistry , fermentation , catalysis , organic chemistry , reaction mechanism , cytochrome p450 , gene
This is the first reported study on the immobilization of living recombinant E scherichia coli cells that overexpress cyclopentanone monooxygenase in polyvinyl alcohol gel particles LentiKats ® . Immobilized cells overexpressing cyclopentanone monooxygenase have been used as a model of biocatalyst for enantioselective B aeyer– V illiger biooxidation of rac‐bicyclo[3.2.0]hept‐2‐en‐6‐one into regioisomeric lactones. This process is useful for the syntheses of cytostatic sarkomycin, several prostaglandins and other biologically active compounds. The original technique for qualitative analysis of enzyme expression within free cells and cells entrapped in L enti K ats ® using SDS ‐ PAGE was developed and used for verification of optimal conditions for the induction of cyclopentanone monooxygenase. Here, we successfully performed six repeated batch Baeyer‐Villiger biooxidations utilizing entrapped cells using 40% (w/v) polyvinyl alcohol gel particles in flasks with baffles. The latter conditions have been found to be the most appropriate achieving optimal oxygen transfer within LentiKats ® . Moreover, immobilized cells retained their catalytic efficiency over six reaction cycles, while the catalytic efficiency of free cells decreased after three reaction cycles. Significance and Impact of the Study Immobilization in polyvinylalcohol gel particles is desirable technique with presumptive impact on industrial applications of recombinant whole‐cell B aeyer– V illiger monooxygenases as biocatalysts for production of bioactive compounds and precursors of potentially new drugs. An original immobilization of cells E . coli with overproduced B aeyer– V illiger monooxygenase improved their stability in repetitive batch biooxidations as compared to free cells. Detected autoinduction of recombinant enzyme in pET 22b+ plays significant role in application of immobilized cells as it may increase specific activity of cells in repetitive use under growing reaction conditions. Original technique for qualitative analysis of enzyme expression within immobilized cells was developed.