Expanding nature's catalytic repertoire to include reactions important in synthetic chemistry will open new opportunities for ‘green’ chemistry and biosynthesis. We demonstrate enzyme-catalyzed insertion of carbenoids into N-H bonds. This type of bond disconnection, which has no counterpart in nature, can be mediated by variants of the cytochrome P450 from Bacillus megaterium. The N-H insertion reaction takes place in water, provides the desired products in 26-83% yield, forms the single addition product exclusively, and does not require slow addition of the diazo component

Engineering enzymes capable of modes of activation unprecedented in nature will increase the range of industrially important molecules that can be synthesized through biocatalysis. However, low activity for a new function is often a limitation in adopting enzymes for preparative scale synthesis, reaction with demanding substrates, or when a natural substrate is also present. By mutating the proximal ligand and other key active site residues of the cytochrome P450 from Bacillus megaterium (P450-BM3), we have engineered a highly active His-ligated variant of P450-BM3, BM3-Hstar, that can be employed for the enantioselective synthesis of the levomilnacipran core. This enzyme catalyzesthe cyclopropanation of N,N-diethyl-2-phenylacrylamide (1) with an estimated initial rate of over 1000 turnovers per minute and can be used under an aerobic environment. Cyclopropanation activity is highly dependent on the electronics of the P450 proximal ligand, which can be used to tune this non-natural enzyme activity.

Engineering enzymes capable of modes of activation unprecedented in nature will increase the range of industrially important molecules that can be synthesized through biocatalysis. However, low activity for a new function is often a limitation in adopting enzymes for preparative scale synthesis, reaction with demanding substrates, or when a natural substrate is also present. By mutating the proximal ligand and other key active site residues of the cytochrome P450 from Bacillus megaterium (P450-BM3), we have engineered a highly active His-ligated variant of P450-BM3, BM3-Hstar, that can be employed for the enantioselective synthesis of the levomilnacipran core. This enzyme catalyzesthe cyclopropanation of N,N-diethyl-2-phenylacrylamide (1) with an estimated initial rate of over 1000 turnovers per minute and can be used under an aerobic environment. Cyclopropanation activity is highly dependent on the electronics of the P450 proximal ligand, which can be used to tune this non-natural enzyme activity. Keywords biocatalysis; cytochrome P450; cyclopropanationEnzymes in nature catalyze only a small subset of industrially-relevant chemical transformations. 1 Thus, increasing the number of activation modes accessible to enzymes is an important goal in biocatalysis, green chemistry, and sustainable synthesis. 2 Drawing an analogy between the mechanism of monooxygenation catalyzed by cytochrome P450 and transition metal-catalyzed carbene insertions, we hypothesized that an iron-carbenoid intermediate could be generated at the enzyme' sheme prosthetic group in the presence of

Extraction techniques utilizing high pH and (bi)carbonate concentrations were evaluated for their efficacy in determining the oxidation state of uranium (U) in reduced sediments collected from Rifle, CO. Differences in dissolved concentrations between oxic and anoxic extractions have been proposed as a means to quantify the U(VI) and U(IV) content of sediments. An additional step was added to anoxic extractions using a strong anion exchange resin to separate dissolved U(IV) and U(VI). X-ray spectroscopy showed that U(IV) in the sediments was present as polymerized precipitates similar to uraninite and/or less ordered U(IV), referred to as non-uraninite U(IV) species associated with biomass (NUSAB). Extractions of sediment containing both uraninite and NUSAB displayed higher dissolved uranium concentrations under oxic than anoxic conditions while extractions of sediment dominated by NUSAB resulted in identical dissolved U concentrations. Dissolved U(IV) was rapidly oxidized under anoxic conditions in all experiments. Uraninite reacted minimally under anoxic conditions but thermodynamic calculations show that its propensity to oxidize is sensitive to solution chemistry and sediment mineralogy. A universal method for quantification of U(IV) and U(VI) in sediments has not yet been developed but the chemical extractions, when combined with solid-phase characterization, have a narrow range of applicability for sediments without U(VI).

Mutation of the axial ligand can be used to tune the carbenoid activity of cytochrome P450 from Bacillus megaterium. In their Communication on , F. Arnold and co‐workers report the engineering of a histidine‐ligated P450 enzyme that shows high catalytic activity for in vivo cyclopropanation using diazo compounds. This enzyme enables the enantioselective synthesis of a key precursor of levomilnacipran under aerobic conditions on a preparative scale. Image: Liang Zong and Yan Liang (L2Molecule.com).