Chiral methyl mandelates are useful synthons in organic transformation and pharmaceutical synthesis. Green synthesis of these valuable compounds by direct C–H activating oxidative hydroxylation has ...attracted keen interest. Described herein is achieving the stereoselective and efficient bio-hydroxylation of methyl 2-phenylacetates to the chiral methyl mandelates by directed evolution of the cytochrome P450DA hydroxylase. In the present study, a new colorimetric high-throughput screening assay was successfully developed based on a dual-enzyme cascade for the engineering of the P450DA's hydroxylation activity. Several beneficial variants with enhanced bio-hydroxylation activity were created by combining random mutagenesis and site-saturated/directed mutagenesis strategies. Whole-cell bio-hydroxylation of various methyl 2-phenylacetates using the best septuplet-mutant P450DA-11 yielded the corresponding chiral methyl mandelates in up to 92% isolated yields and >99% ee.
The cytochrome P450DA hydroxylase was evolved to increase bio-hydroxylation activity toward methyl 2-phenylacetate by developing a new colorimetric high-throughput screening assay. Various chiral methyl mandelates were synthesized via the engineered P450DA-catalyzed stereoselective bio-hydroxylation. Display omitted
•Nano Fe3O4/CMC/SA can exert the advantages of nano Fe3O4 and SA. It has an important research value for immobilized enzyme.•In this study, α-amylase was selected as a simulation object to ...manufacture an immobilized enzyme (SM/CMC/CA/α-Amy).•Immobilization increased 4.67 times to catalytic activity, slowed down release rate and improved reusable performance.•The stabilities of pH, thermal and storage for SM/CMC/CA/α-Amy were improved remarkably.•All these results indicated that SM/CMC/SA was an ideal carrier for immobilizing enzyme.
In order to improve catalytic activity, increase recycling times, reduce use cost for enzyme, superparamagnetic carboxymethyl chitosan/sodium alginate nanosphere (SM/CMC/SA) has been synthesized via an improved hydrothermal method, molecular self-assembly technology, electrostatic interaction and amide linkage. Its mean diameter was 65nm, zeta potential was −36.9mV and BET was 53.8m2/g. α-Amylase was selected as a simulation object to manufacture an immobilized enzyme (SM/CMC/CA/α-Amy), and its catalytic activity, release behavior, reusability and stability were researched. Immobilization increased 4.67 times to catalytic activity, slowed down release rate and improved reusable performance. SM/CMC/CA/α-Amy showed higher activity over a wider pH range, especially in strong acidic and alkaline environments. The thermal stability and storage stability were improved remarkably too. All these results indicated that SM/CMC/SA was an ideal carrier for immobilizing enzyme.
A biocatalytic approach towards a range of 4‐aryloxazolidinones is developed using a halohydrin dehalogenase from Ilumatobacter coccineus (HheG) as biocatalyst. The method is based on the ...HheG‐catalyzed α‐position regioselective ring‐opening of styrene oxide derivatives with cyanate as a nucleophile, producing the corresponding 4‐aryloxazolidinones in moderate to good yields. Synthesis of enantiopure 4‐aryloxazolidinones is also achievable using chiral epoxide materials.
A novel and simple spectrophotometric method is developed for the determination of nitrite in water by using the self-coupling diazotization reagent 2-amino-6-chlorobenzoic acid and UV light ...illumination. The method only involves one reagent and one step, which can be easily applied to the determination of nitrite and tolerates many foreign anions' effects under acidic conditions.
An efficient biocatalytic approach for enantio‐ and regioselective ring‐opening of styrene oxides with cyanate was developed by using the halohydrin dehalogenase HheC from Agrobacterium radiobacter ...AD1, generating the corresponding chiral 5‐aryl‐2‐oxazolidinones in up to 47% yield and 90% ee. Additionally, the origin of enantioselectivity and regioselectivity of the HheC‐catalyzed cyanate‐mediated ring‐opening process was uncovered by single enantiomer bioconversions and molecular docking study.
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•A new function of monoamine oxidase (MAO-5) was found.•Bio-mediated oxidative resolution of racemic 2-substituted indolines was developed.•(R)-2-methyl-indoline and its derivates was ...obtained with high enantioselectivity by biocatalytic process.
Monoamine oxidase strain (MAO-5) from Pseudomonas monteilii ZMU-T01 mediated oxidative resolution of racemic 2-substituted indolines has been successfully described. A series of highly enantioselective 2-substituted indoline derivatives were obtained in up to 50 % conversion and >99 % ee.
A self‐sufficient cytochrome P450 monooxygenase from Deinococcus apachensis (P450DA) was identified and successfully overexpressed in Escherichia coli BL21(DE3). P450DA would be a member of the ...CYP102D subfamily and assigned as CYP102D2 according to the phylogenetic tree and sequence alignment. Purification and characterization of the recombinant P450DA indicated both NADH and NADPH could be used by P450DA as a reducing cofactor. The recombinant E. coli (P450DA) strain was functionally active, showing excellent enantioselectivity for benzylic hydroxylation of methyl 2‐phenylacetate. Further substrate scope studies revealed that P450DA is able to catalyze benzylic hydroxylation of a variety of compounds, affording the corresponding chiral benzylic alcohols in 86–99 % ee and 130–1020 total turnover numbers.
A self‐sufficient cytochrome P450 monooxygenase from Deinococcus apachensis (P450DA) was identified and successfully overexpressed in Escherichia coli BL21(DE3). Further substrate scope studies revealed that P450DA is able to catalyze benzylic hydroxylation of a variety of compounds, affording the corresponding chiral benzylic alcohols in 86–99 % ee and 130–1020 total turnover numbers.
To date, the examples of difunctionalization of alkanes to directly incorporate two functional groups are very limited. In this study, we combined photoorgano redox catalysis and P450 biocatalysts to ...obtain dioxygen-functionalization of α/β-C-H bonds of arylalkanes in a straightforward manner. The synthesis of enantiomerically chiral acyloins through a one-pot two-step photoredox/P450-catalyzed cascade reaction is described. Two P450 mutants with stereocomplementary bio-oxidation were obtained using mutagenesis technology and were able to asymmetrically hydroxylate ketones to acyloins with excellent ee values, which were further proved to be efficient on a wide range of substrates. Moreover, a photoredox synthesis of ketones
in situ
was developed by the direct carbonylation of aromatic methyl C-H bonds and subsequently combined with the aerobic P450-biocatalytic enantioselective hydroxylation of intermediate ketones, thus providing a green and sustainable approach towards optically pure acyloins.
This study has described an efficient, mild, and highly enantioselective one-pot two-step cascade reaction to access chiral α-hydroxyketones from arylalkanes, combining photoredox/P450 enzymatic direct dioxygen-functionalization of arylalkanes α/β C-H bonds.
Halohydrin dehalogenases are usually recognized as strict β-position regioselective enzymes in the nucleophile-mediated ring-opening of epoxides. Here we found the HheG from
exhibited excellent ...α-position regioselectivity in the azide-mediated ring-opening of styrene oxide derivatives 1a-1k, producing the corresponding 2-azido-2-aryl-1-ols 2a-2k with the yields up to 96%.
An efficient strategy for the synthesis of spirocyclic indolizidine derivatives that have a 3,3′‐disubstituted oxindole framework has been described. By using sequential nucleophilic ...C(sp3)‐benzylation and palladium‐catalyzed C(sp2)–H arylation reactions of 3‐pyrrolyloxindoles and 2‐(bromomethyl)aryl bromides, a series of spirooxindole‐3,5′‐pyrrolo2,1‐aisoquinolines were smoothly obtained in up to 92 % yield. A synthetic application of this method was also demonstrated by the transformation of 3u into functionalized compound 4. Moreover, the catalytic cycle of this cascade reaction was proposed on the basis of results of control experiments.
A cooperative base and palladium system was employed for the sequential nucleophilic C(sp3)‐benzylation and C(sp2)–H arylation reactions of 3‐pyrrolyloxindoles and 2‐(bromomethyl)aryl bromides. This protocol was used for the construction of spirocyclic indolizidine derivatives that incorporate a 3,3′‐disubstituted oxindole framework. TFA = trifluoroacetate; DMF = N,N‐dimethylformamide.
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