Cargando…

Biotransformation of Isoflavone Using Enzymatic Reactions

The roles of cytochrome P450 monooxygenases (CYPs) from Streptomyces spp. which are called the “treasure islands” for natural products for medicine and antibiotics are not well understood. Substrate specificity studies on CYPs may give a solution for elucidation of their roles. Based on homology seq...

Descripción completa

Detalles Bibliográficos
Autor principal: Roh, Changhyun
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6270586/
https://www.ncbi.nlm.nih.gov/pubmed/23467013
http://dx.doi.org/10.3390/molecules18033028
_version_ 1783376733148282880
author Roh, Changhyun
author_facet Roh, Changhyun
author_sort Roh, Changhyun
collection PubMed
description The roles of cytochrome P450 monooxygenases (CYPs) from Streptomyces spp. which are called the “treasure islands” for natural products for medicine and antibiotics are not well understood. Substrate specificity studies on CYPs may give a solution for elucidation of their roles. Based on homology sequence information, the CYP105D7 of a soluble cytochrome P450 known as heme protein from Streptomyces avermitilis MA4680 was expressed using the T7 promoter of the bacterial expression vector pET24ma, over-expressed in Escherichia coli system and characterized. An engineered whole cell system for daidzein hydroxylation was constructed using an exogenous electron transport system from ferredoxin reductase (PdR) and ferredoxin (Pdx). Also, an in vitro reaction study showed the purified CYP105D7 enzyme, using NADH-dependent-reducing equivalents of a redox partner from Pseudomonas putida, hydroxylated daidzein at the 3' position of the B ring to produce 7,3,'4' trihydroxyisoflavone. The hydroxylated position was confirmed by GC-MS analysis. The turnover number of the enzyme was 0.69 μmol 7,3,'4'-trihydroxyisoflavone produced per μmol P450 per min. This enzyme CYP105D7 represents a novel type of 3'-hydroxylase for daidzein hydroxylation. A P450 inhibitor such as coumarin significantly (ca.98%) inhibited the daidzein hydroxylation activity.
format Online
Article
Text
id pubmed-6270586
institution National Center for Biotechnology Information
language English
publishDate 2013
publisher MDPI
record_format MEDLINE/PubMed
spelling pubmed-62705862018-12-20 Biotransformation of Isoflavone Using Enzymatic Reactions Roh, Changhyun Molecules Article The roles of cytochrome P450 monooxygenases (CYPs) from Streptomyces spp. which are called the “treasure islands” for natural products for medicine and antibiotics are not well understood. Substrate specificity studies on CYPs may give a solution for elucidation of their roles. Based on homology sequence information, the CYP105D7 of a soluble cytochrome P450 known as heme protein from Streptomyces avermitilis MA4680 was expressed using the T7 promoter of the bacterial expression vector pET24ma, over-expressed in Escherichia coli system and characterized. An engineered whole cell system for daidzein hydroxylation was constructed using an exogenous electron transport system from ferredoxin reductase (PdR) and ferredoxin (Pdx). Also, an in vitro reaction study showed the purified CYP105D7 enzyme, using NADH-dependent-reducing equivalents of a redox partner from Pseudomonas putida, hydroxylated daidzein at the 3' position of the B ring to produce 7,3,'4' trihydroxyisoflavone. The hydroxylated position was confirmed by GC-MS analysis. The turnover number of the enzyme was 0.69 μmol 7,3,'4'-trihydroxyisoflavone produced per μmol P450 per min. This enzyme CYP105D7 represents a novel type of 3'-hydroxylase for daidzein hydroxylation. A P450 inhibitor such as coumarin significantly (ca.98%) inhibited the daidzein hydroxylation activity. MDPI 2013-03-06 /pmc/articles/PMC6270586/ /pubmed/23467013 http://dx.doi.org/10.3390/molecules18033028 Text en © 2013 by the authors; licensee MDPI, Basel, Switzerland. http://creativecommons.org/licenses/by/3.0/ This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/).
spellingShingle Article
Roh, Changhyun
Biotransformation of Isoflavone Using Enzymatic Reactions
title Biotransformation of Isoflavone Using Enzymatic Reactions
title_full Biotransformation of Isoflavone Using Enzymatic Reactions
title_fullStr Biotransformation of Isoflavone Using Enzymatic Reactions
title_full_unstemmed Biotransformation of Isoflavone Using Enzymatic Reactions
title_short Biotransformation of Isoflavone Using Enzymatic Reactions
title_sort biotransformation of isoflavone using enzymatic reactions
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6270586/
https://www.ncbi.nlm.nih.gov/pubmed/23467013
http://dx.doi.org/10.3390/molecules18033028
work_keys_str_mv AT rohchanghyun biotransformationofisoflavoneusingenzymaticreactions