Process limitations of a whole-cell P450 catalyzed reaction using a CYP153A-CPR fusion construct expressed in Escherichia coli

Cytochrome P450s are interesting biocatalysts due to their ability to hydroxylate non-activated hydrocarbons in a selective manner. However, to date only a few P450-catalyzed processes have been implemented in industry due to the difficulty of developing economically feasible processes. In this stud...

Full description

Saved in:

Bibliographic Details
Published in:	Applied microbiology and biotechnology Vol. 100; no. 3; pp. 1197 - 1208
Main Authors:	Lundemo, M. T, Notonier, S, Striedner, G, Hauer, B, Woodley, J. M
Format:	Journal Article
Language:	English
Published:	Berlin/Heidelberg Springer Berlin Heidelberg 01-02-2016 Springer Springer Nature B.V
Subjects:	Bacillus megaterium Bacillus megaterium - enzymology Bacillus megaterium - genetics Bacterial Proteins - chemistry Bacterial Proteins - genetics Bacterial Proteins - metabolism Biocatalysts Biomedical and Life Sciences Biotechnological Products and Process Engineering Biotechnology Catalysis Cytochrome Cytochrome P-450 Cytochrome P-450 Enzyme System - chemistry Cytochrome P-450 Enzyme System - genetics Cytochrome P-450 Enzyme System - metabolism E coli Enzymes Escherichia coli Escherichia coli - genetics Escherichia coli - metabolism Fatty acids Fatty Acids - chemistry Fatty Acids - metabolism Gene Expression Genetic aspects Health aspects heme Hydrocarbons industry Kinetics Life Sciences Marinobacter Marinobacter - enzymology Marinobacter - genetics Marinobacter aquaeolei Microbial Genetics and Genomics Microbiology mutants Observations odors Oxidation Polymers Process engineering Protein Structure, Tertiary Proteins Recombinant Fusion Proteins - chemistry Recombinant Fusion Proteins - genetics Recombinant Fusion Proteins - metabolism Self sufficiency Studies Yeast Europe Process development Bottleneck identification Whole-cell catalysis Cytochrome P450 monooxygenases Inhibition
Online Access:	Get full text
Tags:	Add Tag No Tags, Be the first to tag this record!

Description
Summary:	Cytochrome P450s are interesting biocatalysts due to their ability to hydroxylate non-activated hydrocarbons in a selective manner. However, to date only a few P450-catalyzed processes have been implemented in industry due to the difficulty of developing economically feasible processes. In this study, we have used the CYP153A heme domain from Marinobacter aquaeolei fused to the reductase domain of CYP102A1 from Bacillus megaterium (BM3) expressed in Escherichia coli. This self-sufficient protein chimera CYP153A-CPRBM₃ G307A mutant is able to selectively hydroxylate medium and long chain length fatty acids at the terminal position. ω-Hydroxylated fatty acids can be used in the field of high-end polymers and in the cosmetic and fragrance industry. Here, we have identified the limitations for implementation of a whole-cell P450-catalyzed reaction by characterizing the chosen biocatalyst as well as the reaction system. Despite a well-studied whole-cell P450 catalyst, low activity and poor stability of the artificial fusion construct are the main identified limitations to reach sufficient biocatalyst yield (mass of product/mass of biocatalyst) and space-time yield (volumetric productivity) essential for an economically feasible process. Substrate and product inhibition are also challenges that need to be addressed, and the application of solid substrate is shown to be a promising option to improve the process.
Bibliography:	http://dx.doi.org/10.1007/s00253-015-6999-x ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0175-7598 1432-0614
DOI:	10.1007/s00253-015-6999-x