Structural and Spectroscopic Characterization of Self-Sufficient Cytochrome P450 Fusion Enzymes: CYP102A1 and CYP116B
Year: 2026
Type: preprint
Abstract: Cytochrome P450 (CYP) monooxygenases are heme-thiolate enzymes that catalyze diverse oxidative transformations essential to metabolism, detoxification, and biosynthesis. While most CYPs rely on separate redox partner proteins for electron delivery, a subset-known as self-sufficient or fusion P450s-incorporate the reductase domain(s) directly within a single polypeptide, enabling efficient intramolecular electron transfer and, in some cases, exceptionally high catalytic turnover. This review examines two representative self-sufficient P450 systems, CYP102A1 (P450 BM3) and CYP116B, summarizing spectroscopic evidence (UV-Vis, EPR, and resonance Raman), supported by available crystal structures, to compare how their distinct domain architectures shape heme electronic structure and catalytic mechanism. CYP102A1, a fused FAD/FMN-heme flavocytochrome from Bacillus megaterium, undergoes pronounced substrate-induced conformational rearrangement and low-spin/highspin switching, properties that are further tunable through active-site mutagenesis (e.g., A82F, F87V, F393 variants) and that underlie its status as the most extensively studied and engineered P450 scaffold. In contrast, CYP116B enzymes, which transfer electrons through a defined FMN-[2Fe–2S]-heme pathway, maintain a comparatively rigid, conserved low-spin heme environment that is less dynamically coupled to substrate binding, although engineered variants (e.g., CYP116B5-SOX) demonstrate their potential for peroxide-driven biocatalysis. Together, these comparisons illustrate how differences in fusion architecture translate into distinct strategies for electron transfer and catalytic control, offering complementary design principlesstructural plasticity in BM3 versus a stable redox scaffold in CYP116B-for engineering nextgeneration self-sufficient P450 biocatalysts.
Source: ChemRxiv
Institution: University of Akron
Language: English
Subfield: Inorganic Chemistry
Field: Chemistry
Domain: Physical Sciences
Open Access status: green
OpenAlex ID: W7214667562