Abstract
P450eryF is the only bacterial P450 to show cooperativity of substrate binding and oxidation. However, the studies reported so far have provided evidence only for homotropic cooperativity of P450eryF but not for heterotropic cooperativity. Therefore, oxidation of 7-benzyloxyquinoline (7-BQ) and 1-pyrenebutanol (1-PB) by P450eryF A245T and spectral binding of 9-aminophenanthrene (9-AP) to wild-type P450eryF were investigated in the presence of various effectors. The addition of steroids and flavones caused no stimulation but rather moderate inhibition of 7-BQ or 1-PB oxidation by P450eryF A245T. However, the binding affinity of 9-AP was significantly increased in the presence of androstenedione or α-naphthoflavone (ANF). A comparative study with CYP3A4 revealed a similar increase in the binding affinity of 9-AP for the enzyme at low ANF concentrations but some competition at higher ANF concentrations. These studies, to our knowledge, provide the first report of heterotropic cooperativity in P450eryF as well as spectroscopic evidence for simultaneous presence of two ligand molecules in the CYP3A4 active site.
Footnotes
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↵3 Permanent address: Shanghai Institute of Materia Medica Chinese Academy of Sciences 294 Taiyuan Rd., Shanghai 200031, P. R. China.
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This work was supported by the Robert A. Welch Foundation, Houston, Texas (H1458), Grant GM54995 (JRH) and Center Grant ES06676 from the National Institutes of Health.
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↵2 Recently, Dabrowski et al. (2002)provided spectroscopic evidence for the presence of pyrene dimers in the CYP3A4 active site.
- Abbreviations used are::
- P450
- cytochrome P450
- 9-AP
- 9-aminophenanthrene
- 7-BQ
- 7-benzyloxyquinoline
- 1-PB
- 1-pyrenebutanol
- ANF
- α-naphthoflavone
- Received November 18, 2002.
- Accepted January 8, 2003.
- The American Society for Pharmacology and Experimental Therapeutics
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