Engineered hemoproteins have recently emerged as promising systems for promoting asymmetric cyclopropanations, but variants featuring predictable, complementary stereoselectivity in these reactions have remained elusive. In this study, a rationally driven strategy was implemented and applied to engineer myoglobin variants capable of providing access to 1-carboxy-2-aryl-cyclopropanes with high trans-(1R,2R) selectivity and catalytic activity. The stereoselectivity of these cyclopropanation biocatalysts complements that of trans-(1S,2S)-selective variants developed here and previously. In combination with whole-cell biotransformations, these stereocomplementary biocatalysts enabled the multigram synthesis of the chiral cyclopropane core of four drugs (Tranylcypromine, Tasimelteon, Ticagrelor, TRPV1 inhibitor 24) in high yield and with excellent diastereo- and enantioselectivity (98–99.9% de; 96–99.9% ee). These biocatalytic strategies outperform currently available methods to produce these drugs.

Novel bisbenzimidazole inhibitors of bacterial type IA topoisomerase are of interest for the development of new antibacterial agents that are impacted by target-mediated cross resistance with fluoroquinolones. The present study demonstrates the successful synthesis and evaluation of bisbenzimidazole analogues as Escherichia coli topoisomerase IA inhibitors. 5-(4-Propylpiperazin-1-yl)-2-[2'-(4-ethoxyphenyl)-5'-benzimidazolyl]benzimidazole (12b) showed significant relaxation inhibition activity against EcTopo 1A (IC50 = 2 ± 0.005 μM) and a tendency to chelate metal ion. Interestingly, these compounds did not show significant inhibition of E. coli DNA gyrase and hTop 1 even up to 100 μM. Compound 12b has shown lowest MIC against E. coli strains among 24 compounds evaluated. The binding affinity constant and binding free energy of 12b with EcTopo 1A was observed 6.8 × 10(6) M(-1) and -10.84 kcal mol(-1) from isothermal titration calorimetry (ITC), respectively. In vivo mouse systemic infection and neutropenic thigh model experimental results confirmed the therapeutic efficacy of 12b, suggesting further development of this class of compounds as antibacterial agents.

Escherichia coli strains belonging to diverse pathotypes have increasingly been recognized as a major public health concern. The β-lactam antibiotics have been used successfully to treat infections caused by pathogenic E. coli. However, currently, the utility of β-lactams is being challenged severely by a large number of hydrolytic enzymes – the β-lactamases expressed by bacteria. The menace is further compounded by the highly flexible genome of E. coli, and propensity of resistance dissemination through horizontal gene transfer and clonal spread. Successful management of infections caused by such resistant strains requires an understanding of the diversity of β-lactamases, their unambiguous detection, and molecular mechanisms underlying their expression and spread with regard to the most relevant information about individual bacterial species. Thus, this review comprises first such effort in this direction for E. coli, a bacterial species known to be associated with production of diverse classes of β-lactamases. The review also highlights the role of commensal E. coli as a potential but under-estimated reservoir of β-lactamases-encoding genes.

The first example of a biocatalytic [2,3]-sigmatropic rearrangement reaction involving allylic sulfides and diazo reagents (Doyle-Kirmse reaction) is reported. Engineered variants of sperm whale myoglobin catalyze this synthetically valuable C–C bond forming transformation with high efficiency and product conversions across a variety of sulfide substrates (i.e., aryl-, benzyl-, and alkyl-substituted allylic sulfides) and α-diazo esters. Moreover, the scope of this myoglobin-mediated transformation could be extended to the conversion of propargylic sulfides to give substituted allenes. Active site mutations proved effective toward enhancing the catalytic efficiency of the hemoprotein in these reactions as well as modulating its enantioselectivity, resulting in the identification of a myoglobin variant, Mb(L29S, H64V, V68F), capable of mediating asymmetric Doyle-Kirmse reactions with an enantiomeric excess up to 71%. This work extends the toolbox of currently available biocatalytic strategies for realizing the asymmetric formation of carbon–carbon bonds.

Hemoproteins have recently emerged as promising biocatalysts for promoting a variety of carbene transfer reactions including cyclopropanation and Y-H insertion (Y = N, S, Si, B). For these and synthetic carbene transfer catalysts alike, achieving high chemoselectivity toward cyclopropanation in olefin substrates bearing unprotected Y-H groups has proven remarkably challenging due to competition from the more facile carbene Y-H insertion reaction. In this report, we describe the development of a novel artificial metalloenzyme based on an engineered myoglobin incorporating a serine-ligated Co-porphyrin cofactor that is capable of offering high selectivity toward olefin cyclopropanation over N-H and Si-H insertion. Intramolecular competition experiments revealed a distinct and dramatically altered chemoselectivity of the Mb(H64V,V68A,H93S)[Co(ppIX)] variant in carbene transfer reactions compared to myoglobin-based variants containing the native histidine-ligated heme cofactor or other metal/proximal ligand substitutions. These studies highlight the functional plasticity of myoglobin as a "carbene transferase" and illustrate how modulation of the cofactor environment within this metalloprotein scaffold represents a valuable strategy for accessing carbene transfer reactivity not exhibited by naturally occurring hemoproteins or transition metal catalysts.

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Quinolone and β-lactam antibiotics constitute major mainstay of treatment against infections caused by pathogenic Escherichia coli. Presence of E. coli strains expressing co-resistance to both these antibiotic classes in urban aquatic environments which are consistently being used for various anthropogenic activities represents a serious public health concern. From a heterogeneous collection of 61 E. coli strains isolated from the river Yamuna traversing through the National Capital Territory of Delhi (India), those harboring blaCTX-M-15 (n = 10) or blaCMY-42 (n = 2) were investigated for co-resistance to quinolones and the molecular mechanisms thereof. Resistance was primarily attributed to amino acid substitutions in the quinolone resistance-determining regions (QRDRs) of GyrA (S83L ± D87N) and ParC (S80I ± E84K). One of the E. coli strains, viz., IPE, also carried substitutions in GyrB and ParE at positions Ser492→Asn and Ser458→Ala, respectively. The phenotypically susceptible strains nevertheless carried plasmid-mediated quinolone resistance (PMQR) gene, viz., qnrS, which showed co-transfer to the recipient quinolone-sensitive E. coli J53 along with the genes encoding β-lactamases and led to increase in minimal inhibitory concentrations of quinolone antibiotics. To the best of our knowledge, this represents first report of molecular characterization of quinolone co-resistance in E. coli harboring genes for ESBLs or AmpC β-lactamases from a natural aquatic environment of India. The study warrants true appreciation of the potential of urban aquatic environments in the emergence and spread of multi-drug resistance and underscores the need to characterize resistance genetic elements vis-à-vis their public health implications, irrespective of apparent phenotypic resistance.