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Cited by 44Steroid Chemistry and BiochemistryLipid Membrane Structure and BehaviorPharmacogenetics and Drug Metabolism

Interaction of human CYP17 (P-45017α, 17α-hydroxylase-17,20 lyase) with cytochrome b5: importance of the orientation of the hydrophobic domain of cytochrome b5

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LEE-ROBICHAUD, Peter, KADERBHAI, Mustak A., KADERBHAI, Naheed, WRIGHT, J. Neville, AKHTAR, Muhammad·1997·Biochemical Journal
DOI 10.1042/bj3210857PMC PMC1218145
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Abstract

Human CYP17 (P-45017α, 17α-hydroxylase-17,20-lyase)-catalysed side-chain cleavage of 17α-hydroxyprogestogens into androgens is greatly dependent on the presence of cytochrome b5. The native form of cytochrome b5 is composed of a globular core, residues 1Ő98, followed by a membrane insertable C-terminal tail, residues 99Ő133. In the present study the abilities of five different forms of cytochrome b5 to support the side-chain cleavage activity of CYP17 were compared. The five derivatives were: the native pig cytochrome b5 (native pig), its genetically engineered rat counterpart (coreŐtail), the soluble core form of the latter (core), the core with the secretory signal sequence of alkaline phosphatase appended to its N-terminal (signalŐcore) and the latter containing the C-terminal tail of the native rat protein (signalŐcoreŐtail). When examined by Edman degradation and MS, the engineered proteins were shown to have the expected N-terminal amino acid sequences and molecular masses. The native pig was found to be acetylated at the N-terminal. The native pig and coreŐtail enzymes were equally efficient at enhancing the side-chain cleavage activity of human CYP17 and the signalŐcoreŐtail was 55% as efficient. The core and signalŐcore constructs were completely inactive in the aforementioned reaction. All the five derivatives were reduced to varying degrees by NADPH:cytochrome P-450 (NADPH-P450) reductase and the relative efficiencies of this reduction were reminiscent of the behaviour of these derivatives in supporting the side-chain cleavage reaction. In the side-chain cleavage assay, however, NADPH-P450 reductase was used in large excess so that the reduction of cytochrome b5 derivatives was not rate-limiting. The results highlight that productive interaction between cytochrome b5 and CYP17 is governed not only by the presence of a membrane insertable hydrophobic region on the cytochrome b5 but also by its defined spatial orientation at the C-terminal.

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  • Genetic and biochemical studies have implicated basic residues in P450c17, including R347, R358, and perhaps R449 and K89, as important for its interaction with cytochrome b5, whereas E48 and E49 of cytochrome b5 are required for high 17,20-lyase activity.

    “Genetic and biochemical studies have implicated basic residues in P450c17, including R347, R358, and perhaps R449 and K89, as important for its interaction with cytochrome b 5 (240,280,348), whereas E48 and E49 of cytochrome b 5 are required for high 17,20-lyase activity (349).”

    From The Molecular Biology, Biochemistry, and Physiology of Human Steroidogenesis and Its Disorders (2011) · cites this paper as [348]

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  • The C-terminal membrane-spanning helix (exiting the core 2 domain) is required to stimulate the 17,20-lyase activity of human P450c17, but the signal peptide is not.

    “Finally, the C-terminal membrane-spanning helix (exiting the core 2 domain) is required to stimulate the 17,20-lyase activity of human P450c17, but the signal peptide is not (348).”

    From The Molecular Biology, Biochemistry, and Physiology of Human Steroidogenesis and Its Disorders (2011) · cites this paper as [348]

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Research works printed but not linked: 2

  1. [24]24. Akhtar, M., Corina, D. L., Miller, S. L., Shyadehi, A. Z. and Wright, J. N. (1994) J. Chem. Soc., Perkin Trans. 1 263-267
  2. [33]33. Spatz, L. and Strittmatter, P. (1971) Proc. Natl. Acad. Sci. U.S.A. 68, 1042-1046

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Authors
LEE-ROBICHAUD, P., KADERBHAI, M. A., KADERBHAI, N., WRIGHT, J. N., & AKHTAR, M.
Affiliations
  • University of Southampton (GB)
  • Aberystwyth University (GB)
Corresponding author
AKHTAR, Muhammad
ORCID
AKHTAR, Muhammad
Venue
Biochemical Journal
Publisher
Portland Press Ltd.
Published
1997-02-01
Volume
321
Issue
3
Pages
857-864
ISSN
  • 0264-6021 (ISSN-L)
  • 1470-8728
DOI
10.1042/bj3210857
PMC
PMC1218145
OpenAlex
W1907523994
Semantic Scholar
fb77c096dbf4ef0a967fe814e6f4d7c388721263
Cited by
44
References
42
Citations by year
2022: 1 · 2019: 1 · 2016: 1 · 2015: 4 · 2014: 5 · 2013: 3 · 2012: 2
FWCI
0.95
Type
journal article
Language
en
Open access
Locations
  • Biochemical Journal (publishedVersion)
  • PubMed Central (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • Wellcome Trust
  • Biotechnology and Biological Sciences Research Council
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Steroid Chemistry and Biochemistry
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Lipid Membrane Structure and Behavior
  • Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmacology › Pharmacogenetics and Drug Metabolism
Keywords
CYP17, cytochrome b5, 17α-hydroxylase/17,20-lyase, side-chain cleavage, androgen production, protein-protein interaction, NADPH-cytochrome P450 reductase, N-terminal acetylation
MeSH terms
Acetaldehyde, Amino Acid Sequence, Animals, Cytochrome P-450 Enzyme System, Humans, Membrane Proteins, Models, Biological, Molecular Sequence Data, Peptide Fragments, Recombinant Proteins, Steroid 17-alpha-Hydroxylase, Molecular Structure, Cytochromes b5, Sequence Analysis, Protein Sorting Signals, Rats

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