BIOspektrum, Journal Year: 2024, Volume and Issue: 30(5), P. 590 - 592
Published: Sept. 1, 2024
Language: Английский
Methods in enzymology on CD-ROM/Methods in enzymology, Journal Year: 2025, Volume and Issue: unknown
Published: Jan. 1, 2025
Language: Английский
Citations
0
ACS Catalysis, Journal Year: 2025, Volume and Issue: unknown, P. 4160 - 4171
Published: Feb. 23, 2025
Language: Английский
Citations
0
Journal of Agricultural and Food Chemistry, Journal Year: 2025, Volume and Issue: unknown
Published: March 20, 2025
Cytochrome P450 enzymes (P450s) are promising candidates for the biosynthesis of 25-hydroxyvitamin D3 (25(OH)VD3). However, their industrial application is limited by challenges, such as low stability, inefficient catalysis, and uncoupling reactions. The construction self-sufficient P450s offers a strategic solution to these limitations, but requires linker optimization regulate interdomain conformational dynamics. In this study, we integrated whole-cell biocatalyst screening with systematic reaction conditions, including cosolvents, cell concentrations, plasmid selection, enhance catalytic performance. Under optimized heme domain Vdh-K1 achieved 91.6% conversion efficiency was subsequently selected chimeric enzyme assembly. By employing local energetic frustration analysis evaluate protein flexibility allosteric dynamics, identified variants highly frustrated linkers. optimal variant, VK1-CYP116B46-L21, exhibited improved thermostability, activity, coupling efficiency, achieving yield 4.89 mM (1.96 g/L) 25(OH)VD3 in Escherichia coli catalysis─the highest reported date. This work underscores utility computational refining dynamics multidomain establishes scalable, cost-effective framework advance systems high-value compounds.
Language: Английский
Citations
0
Journal of Agricultural and Food Chemistry, Journal Year: 2025, Volume and Issue: unknown
Published: April 16, 2025
25-hydroxyvitamin D3 (25(OH)VD3) is an important daily nutritional supplement, and direct enzymatic C25 hydroxylation of vitamin (VD3) to 25(OH)VD3 a green sustainable method. However, the low catalytic activity electron transfer efficiency redox partners cytochrome P450 limited its production. Here, structure-guided semirational design CYP109E1 led mutant CYP109E1M2, which showed 2-fold increase in production compared wild-type. Furthermore, novel reductase domain BmRD was first employed construct fusion protein with CYP109E1M2. Notably, truncating only two amino acids at N-terminus generated Chimera-2, resulting 38.5% Under optimized conditions, engineered strain Escherichia coli 03-2 produced 491.3 mg/L conversion 49.0% space-time yield 61.4 mg/(L·h). This work demonstrates modification optimization potential lays theoretical foundation for industrialization biosynthesis.
Language: Английский
Citations
0
Chemistry - A European Journal, Journal Year: 2024, Volume and Issue: unknown
Published: Aug. 22, 2024
Abstract Vitamin D deficiency affects nearly half the population, with many requiring or opting for supplements vitamin 3 (VD ), precursor of (1α,25‐dihydroxyVD ). 25‐HydroxyVD , circulating form D, is a more effective supplement than VD but its synthesis complex. We report here engineering cytochrome P450 BM3 (CYP102A1) selective oxidation to 25‐hydroxyVD . Long‐range effects substrate‐channel mutation Glu435Ile promoted binding side chain close heme, enhancing activity that reached 6.62 g isolated from 1‐litre scale reaction (69.1 % yield; space‐time‐yield 331 mg/L/h).
Language: Английский
Citations
2
BIOspektrum, Journal Year: 2024, Volume and Issue: 30(5), P. 590 - 592
Published: Sept. 1, 2024
Language: Английский
Citations
0