A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET).
Effector-induced allosteric transitions in cytochrome P450 3A4 (CYP3A4) were investigated by luminescence resonance energy transfer (LRET) between two SH-reactive probes attached to various pairs of distantly located cysteine residues, namely the double-cysteine mutants CYP3A4(C64/C468), CYP3A4(C377...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2013-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC3871636?pdf=render |
| _version_ | 1819038782986387456 |
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| author | Elena V Sineva Jessica A O Rumfeldt James R Halpert Dmitri R Davydov |
| author_facet | Elena V Sineva Jessica A O Rumfeldt James R Halpert Dmitri R Davydov |
| author_sort | Elena V Sineva |
| collection | DOAJ |
| description | Effector-induced allosteric transitions in cytochrome P450 3A4 (CYP3A4) were investigated by luminescence resonance energy transfer (LRET) between two SH-reactive probes attached to various pairs of distantly located cysteine residues, namely the double-cysteine mutants CYP3A4(C64/C468), CYP3A4(C377/C468) and CYP3A4(C64/C121). Successive equimolar labeling of these proteins with the phosphorescent probe erythrosine iodoacetamide (donor) and the near-infrared fluorophore DY-731 maleimide (acceptor) allowed us to establish donor/acceptor pairs sensitive to conformational motions. The interactions of all three double-labeled mutants with the allosteric activators α-naphthoflavone and testosterone resulted in an increase in the distance between the probes. A similar effect was elicited by cholesterol. These changes in distance vary from 1.3 to 8.5 Å, depending on the position of the donor/acceptor pair and the nature of the effector. In contrast, the changes in the interprobe distance caused by such substrates as bromocriptine or 1-pyrenebutanol were only marginal. Our results provide a decisive support to the paradigm of allosteric modulation of CYP3A4 and indicate that the conformational transition caused by allosteric effectors increases the spatial separation between the beta-domain of the enzyme (bearing residues Cys64 and Cys377) and the alpha-domain, where Cys121 and Cys468 are located. |
| first_indexed | 2024-12-21T08:42:47Z |
| format | Article |
| id | doaj.art-264727140a7b458488f802a80190e07d |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-21T08:42:47Z |
| publishDate | 2013-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-264727140a7b458488f802a80190e07d2022-12-21T19:09:54ZengPublic Library of Science (PLoS)PLoS ONE1932-62032013-01-01812e8389810.1371/journal.pone.0083898A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET).Elena V SinevaJessica A O RumfeldtJames R HalpertDmitri R DavydovEffector-induced allosteric transitions in cytochrome P450 3A4 (CYP3A4) were investigated by luminescence resonance energy transfer (LRET) between two SH-reactive probes attached to various pairs of distantly located cysteine residues, namely the double-cysteine mutants CYP3A4(C64/C468), CYP3A4(C377/C468) and CYP3A4(C64/C121). Successive equimolar labeling of these proteins with the phosphorescent probe erythrosine iodoacetamide (donor) and the near-infrared fluorophore DY-731 maleimide (acceptor) allowed us to establish donor/acceptor pairs sensitive to conformational motions. The interactions of all three double-labeled mutants with the allosteric activators α-naphthoflavone and testosterone resulted in an increase in the distance between the probes. A similar effect was elicited by cholesterol. These changes in distance vary from 1.3 to 8.5 Å, depending on the position of the donor/acceptor pair and the nature of the effector. In contrast, the changes in the interprobe distance caused by such substrates as bromocriptine or 1-pyrenebutanol were only marginal. Our results provide a decisive support to the paradigm of allosteric modulation of CYP3A4 and indicate that the conformational transition caused by allosteric effectors increases the spatial separation between the beta-domain of the enzyme (bearing residues Cys64 and Cys377) and the alpha-domain, where Cys121 and Cys468 are located.http://europepmc.org/articles/PMC3871636?pdf=render |
| spellingShingle | Elena V Sineva Jessica A O Rumfeldt James R Halpert Dmitri R Davydov A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET). PLoS ONE |
| title | A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET). |
| title_full | A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET). |
| title_fullStr | A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET). |
| title_full_unstemmed | A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET). |
| title_short | A large-scale allosteric transition in cytochrome P450 3A4 revealed by luminescence resonance energy transfer (LRET). |
| title_sort | large scale allosteric transition in cytochrome p450 3a4 revealed by luminescence resonance energy transfer lret |
| url | http://europepmc.org/articles/PMC3871636?pdf=render |
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