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The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450
Cytochrome P450 reductase (CYPOR) provides electrons to all human microsomal cytochrome P450s (cyt P450s). The length and sequence of the “140s” FMN binding loop of CYPOR has been shown to be a key determinant of its redox potential and activity with cyt P450s. Shortening the “140s loop” by deleting...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8508823/ https://www.ncbi.nlm.nih.gov/pubmed/34638963 http://dx.doi.org/10.3390/ijms221910625 |
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author | Rwere, Freeborn Im, Sangchoul Waskell, Lucy |
author_facet | Rwere, Freeborn Im, Sangchoul Waskell, Lucy |
author_sort | Rwere, Freeborn |
collection | PubMed |
description | Cytochrome P450 reductase (CYPOR) provides electrons to all human microsomal cytochrome P450s (cyt P450s). The length and sequence of the “140s” FMN binding loop of CYPOR has been shown to be a key determinant of its redox potential and activity with cyt P450s. Shortening the “140s loop” by deleting glycine-141(ΔGly141) and by engineering a second mutant that mimics flavo-cytochrome P450 BM3 (ΔGly141/Glu142Asn) resulted in mutants that formed an unstable anionic semiquinone. In an attempt to understand the molecular basis of the inability of these mutants to support activity with cyt P450, we expressed, purified, and determined their ability to reduce ferric P450. Our results showed that the ΔGly141 mutant with a very mobile loop only reduced ~7% of cyt P450 with a rate similar to that of the wild type. On the other hand, the more stable loop in the ΔGly141/Glu142Asn mutant allowed for ~55% of the cyt P450 to be reduced ~60% faster than the wild type. Our results reveal that the poor activity of the ΔGly141 mutant is primarily accounted for by its markedly diminished ability to reduce ferric cyt P450. In contrast, the poor activity of the ΔGly141/Glu142Asn mutant is presumably a consequence of the altered structure and mobility of the “140s loop”. |
format | Online Article Text |
id | pubmed-8508823 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-85088232021-10-13 The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450 Rwere, Freeborn Im, Sangchoul Waskell, Lucy Int J Mol Sci Communication Cytochrome P450 reductase (CYPOR) provides electrons to all human microsomal cytochrome P450s (cyt P450s). The length and sequence of the “140s” FMN binding loop of CYPOR has been shown to be a key determinant of its redox potential and activity with cyt P450s. Shortening the “140s loop” by deleting glycine-141(ΔGly141) and by engineering a second mutant that mimics flavo-cytochrome P450 BM3 (ΔGly141/Glu142Asn) resulted in mutants that formed an unstable anionic semiquinone. In an attempt to understand the molecular basis of the inability of these mutants to support activity with cyt P450, we expressed, purified, and determined their ability to reduce ferric P450. Our results showed that the ΔGly141 mutant with a very mobile loop only reduced ~7% of cyt P450 with a rate similar to that of the wild type. On the other hand, the more stable loop in the ΔGly141/Glu142Asn mutant allowed for ~55% of the cyt P450 to be reduced ~60% faster than the wild type. Our results reveal that the poor activity of the ΔGly141 mutant is primarily accounted for by its markedly diminished ability to reduce ferric cyt P450. In contrast, the poor activity of the ΔGly141/Glu142Asn mutant is presumably a consequence of the altered structure and mobility of the “140s loop”. MDPI 2021-09-30 /pmc/articles/PMC8508823/ /pubmed/34638963 http://dx.doi.org/10.3390/ijms221910625 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Communication Rwere, Freeborn Im, Sangchoul Waskell, Lucy The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450 |
title | The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450 |
title_full | The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450 |
title_fullStr | The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450 |
title_full_unstemmed | The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450 |
title_short | The FMN “140s Loop” of Cytochrome P450 Reductase Controls Electron Transfer to Cytochrome P450 |
title_sort | fmn “140s loop” of cytochrome p450 reductase controls electron transfer to cytochrome p450 |
topic | Communication |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8508823/ https://www.ncbi.nlm.nih.gov/pubmed/34638963 http://dx.doi.org/10.3390/ijms221910625 |
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