Regular ArticleExpression in Escherichia coli of Functional Cytochrome P450c17 Lacking Its Hydrophobic Amino-Terminal Signal Anchor
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Generation of human steroidogenic cytochrome P450 enzymes for structural and functional characterization
2023, Methods in EnzymologyInfluence of Transmembrane Helix Mutations on Cytochrome P450-Membrane Interactions and Function
2019, Biophysical JournalEfficiency of the sulfate pathway in comparison to the Δ4- and Δ5-pathway of steroidogenesis in the porcine testis
2018, Journal of Steroid Biochemistry and Molecular BiologyCitation Excerpt :The cDNA of porcine CYP17A1 was synthesized (Thermo Fisher GeneArt, Regensburg, Germany) and cloned through the restriction sites NdeI and BamHI into the pET-17b plasmid. The N-terminal hydrophobic anchor of the amino acid sequence of CYP17A1 was deleted in order to increase the solubility [52] and the C-terminus was extended by a hexa-histidine tag to facilitate purification. The cDNAs of bovine b5 (isoform CYB5A) and bovine CPR were obtained through amplification from a cDNA library of bovine liver (Zyagen, California, USA) and cloned into the pET-17b plasmid through the restriction sites NdeI and BamHI, as described [57].
Functionalized poly(3-hydroxybutyric acid) bodies as new in vitro biocatalysts
2018, Biochimica et Biophysica Acta - Proteins and ProteomicsEffects of membrane mimetics on cytochrome P450-cytochrome b5 interactions characterized by NMR spectroscopy
2015, Journal of Biological ChemistryCitation Excerpt :The possible reasons for the capability of isotropic bicelles to enhance cytb5-CYP2B4 interactions could be that 1) both proteins are anchored into the membrane through their TM domains so that they are in close proximity to each other, or 2) the membrane directly interacts with the non-TM part of CYP2B4, the tr-CYP2B4, which assists in orienting the protein for optimal contact with cytb5. Although a large body of evidence has been reported to support the hypothesis that interactions between the membrane surface and tr-P450s (lacking the TM domain) exist (17–22, 62–66), it remains vague what role this potential interaction plays in the complex formation between cytb5 and P450. Therefore, it is of great interest to explore the interactions between cytb5 and the tr-P450 under the influence of a membrane environment.
A steroidogenic pathway for sulfonated steroids: The metabolism of pregnenolone sulfate
2014, Journal of Steroid Biochemistry and Molecular BiologyCitation Excerpt :CYP17A1 cDNA was kindly provided by Prof. M. Waterman (Vanderbilt University, Nashville, USA). The amino acid sequence of CYP17A1 is lacking its N-terminal hydrophobic anchor [29] and is extended at the C-terminus by a hexa-histidine-tag to facilitate purification. The CPR amino acid sequence is extended at the C-terminus by three glycines and six histidines and at the N-terminus has a lack of the first 27 amino acids [30].