μ-Oxo-bridged diiron(iii) complexes of tripodal 4N ligands as catalysts for alkane hydroxylation reaction using m-CPBA as an oxidant: substrate vs. self hydroxylation

The ligand stereoelectronic effect of diiron(iii) complexes determines the efficiency and selectivity of catalytic alkane hydroxylation with m-CPBA as an oxidant.

Enhancing catalytic alkane hydroxylation by tuning the outer coordination sphere in a heme-containing metal–organic framework

Removing labile protons via acetylation of the structural nodes in a metal–organic framework enhances catalysis at the heme linkers.

Efficient alkane hydroxylation catalysis of nickel(ii) complexes with oxazoline donor containing tripodal tetradentate ligands

Nickel(ii) complexes with oxazoline-based tripodal ligands are efficient catalysts for alkane oxidation, and reaction rates and oxidizing position selectivity are controlled by the structural properties of the substituent groups on the ligands.

Deconstruction of the CYP153A6 Alkane Hydroxylase System: Limitations and Optimization of In Vitro Alkane Hydroxylation

Some of the most promising results for bacterial alkane hydroxylation to alcohols have been obtained with the cytochrome P450 monooxygenase CYP153A6. CYP153A6 belongs to the class I CYPs and is generally expressed from an operon that also encodes the ferredoxin (Fdx) and ferredoxin reductase (FdR) which transfer electrons to CYP153A6. In this study, purified enzymes (CYP, Fdx, FdR and dehydrogenases for cofactor regeneration) were used to deconstruct the CYP153A6 system into its separate components, to investigate the factors limiting octane hydroxylation in vitro. Proteins in the cytoplasm (cell-free extract) were found to better enhance and stabilize hydroxylase activity compared to bovine serum albumin (BSA) and catalase. Optimization of the CYP:Fdx:FdR ratio also significantly improved both turnover frequencies (TFs) and total turnover numbers (TTNs) with the ratio of 1:1:60 giving the highest values of 3872 h−1 and 45,828 moloctanol molCYP−1, respectively. Choice and concentration of dehydrogenase for cofactor regeneration also significantly influenced the reaction. Glucose dehydrogenase concentrations had to be as low as possible to avoid fast acidification of the reaction medium, which in the extreme caused precipitation of the CYP and other proteins. Cofactor regeneration based on glycerol failed, likely due to accumulation of dihydroxyacetone. Scaling the reactions up from 1 mL in vials to 60 mL in shake flasks and 120 mL in bioreactors showed that mixing and shear forces will be important obstacles to overcome in preparative scale reactions.

Production of 1-Dodecanol, 1-Tetradecanol, and 1,12-Dodecanediol through Whole-Cell Biotransformation inEscherichia coli

ABSTRACTMedium- and long-chain 1-alkanol and α,ω-alkanediols are used in personal care products, in industrial lubricants, and as precursors for polymers synthesized for medical applications. The industrial production of α,ω-alkanediols by alkane hydroxylation primarily occurs at high temperature and pressure using heavy metal catalysts. However, bioproduction has recently emerged as a more economical and environmentally friendly alternative. Among alkane monooxygenases, CYP153A fromMarinobacter aquaeoleiVT8 (CYP153AM.aq; the strain is also known asMarinobacter hydrocarbonoclasticusVT8) possesses low overoxidation activity and high regioselectivity and thus has great potential for use in terminal hydroxylation. However, the application of CYP153AM.aqis limited because it is encoded by a dysfunctional operon. In this study, we demonstrated that the operon regulator AlkRM.aqis functional, can be induced by alkanes of various lengths, and does not suffer from product inhibition. Additionally, we identified a transposon insertion in the CYP153AM.aqoperon. When the transposon was removed, the expression of the operon genes could be induced by alkanes, and the alkanes could then be oxyfunctionalized by the resulting proteins. To increase the accessibility of medium- and long-chain alkanes, we coexpressed a tunable alkane facilitator (AlkL) fromPseudomonas putidaGPo1. Using a recombinantEscherichia colistrain, we produced 1.5 g/liter 1-dodecanol in 20 h and 2 g/liter 1-tetradecanol in 50 h by adding dodecane and tetradecane, respectively. Furthermore, in 68 h, we generated 3.76 g/liter of 1,12-dodecanediol by adding a dodecane–1-dodecanol substrate mixture. This study reports a very efficient method of producing C12/C14alkanols and C121,12-alkanediol by whole-cell biotransformation.IMPORTANCETo produce terminally hydroxylated medium- to long-chain alkane compounds by whole-cell biotransformation, substrate permeability, enzymatic activity, and the control of overoxidability should be considered. Due to difficulties in production, small amounts of 1-dodecanol, 1-tetradecanol, and 1,12-dodecanediol are typically produced. In this study, we identified an alkane-inducible monooxygenase operon that can efficiently catalyze the conversion of alkane to 1-alkanol with no detection of the overoxidation product. By coexpressing an alkane membrane facilitator, high levels of 1-dodecanol, 1-tetradecanol, and 1,12-dodecanediol could be generated. This study is significant for the bioproduction of medium- and long-chain 1-alkanol and α,ω-alkanediols.