Modifying the Basicity of the dNTP Leaving Group Modulates Precatalytic Conformational Changes of DNA Polymerase β.

The catalytic function of DNA polymerase β (pol β) fulfills the gap-filling requirement of the base excision DNA repair pathway by incorporating a single nucleotide into a gapped DNA substrate resulting from the removal of damaged DNA bases. Most importantly, pol β can select the correct nucleotide from a pool of similarly structured nucleotides to incorporate into DNA in order to prevent the accumulation of mutations in the genome. Pol β is likely to employ various mechanisms for substrate selection.

Azetidinimines as a novel series of non-covalent broad-spectrum inhibitors of β-lactamases with submicromolar activities against carbapenemases KPC-2 (class A), NDM-1 (class B) and OXA-48 (class D).

The occurrence of resistances in Gram negative bacteria is steadily increasing to reach extremely worrying levels and one of the main causes of resistance is the massive spread of very efficient β-lactamases which render most β-lactam antibiotics useless. Herein, we report the development of a series of imino-analogues of β-lactams (namely azetidinimines) as efficient non-covalent inhibitors of β-lactamases. Despite the structural and mechanistic differences between serine-β-lactamases KPC-2 and OXA-48 and metallo-β-lactamase NDM-1, all three enzymes can be inhibited at a submicromolar level by compound 7dfm, which can also repotentiate imipenem against a resistant strain of Escherichia coli expressing NDM-1.

Completing the β,γ-CXY-dNTP Stereochemical Probe Toolkit: Synthetic Access to the dCTP Diastereomers and P and F NMR Correlations with Absolute Configurations.

Nucleoside 5'-triphosphate (dNTP) analogues in which the β,γ-oxygen is mimicked by a CXY group (β,γ-CXY-dNTPs) have provided information about DNA polymerase catalysis and fidelity. Definition of CXY stereochemistry is important to elucidate precise binding modes. We previously reported the ()- and ()-β,γ-CHX-dGTP diastereomers (X = F, Cl), prepared via P,C-dimorpholinamide CHCl (, ) and CHF (, ) bisphosphonates (BPs) equipped with an ()-mandelic acid as a chiral auxiliary, with final deprotection using H/Pd.

New Chirally Modified Bisphosphonates for Synthesis of Individual Beta,Gamma-CHX-Deoxynucleotide Diastereomers.

Individual diastereomers of CXY bisphosphonate analogues of dNTPs or NTPs are useful chemical stereoprobes to investigate interactions within the chiral active site environment of enzymes such as polymerases and kinases. We previously reported synthetic access to β,γ-CHX-dGTPs (X = F or Cl) via a bisphosphonate synthon with an (R)-methyl mandelate auxiliary and have extended this approach to dTTP and dATP analogues. As removal of the chiral auxiliary by (Pd/C) hydrogenolysis is incompatible with the cytosine heterocycle and also with X = Br, we have now designed bisphosphonate synthons using (R)-(+)-α-ethylbenzylamine or methyl (R)-(-)-phenylglycine auxiliaries and equipped with an o-nitrobenzyl ester protecting group allowing photochemical deprotection.

Probing DNA Base-Dependent Leaving Group Kinetic Effects on the DNA Polymerase Transition State.

We examine the DNA polymerase β (pol β) transition state (TS) from a leaving group pre-steady-state kinetics perspective by measuring the rate of incorporation of dNTPs and corresponding novel β,γ-CXY-dNTP analogues, including individual β,γ-CHF and -CHCl diastereomers with defined stereochemistry at the bridging carbon, during the formation of right (R) and wrong (W) base pairs. Brønsted plots of log k versus p K of the leaving group bisphosphonic acids are used to interrogate the effects of the base identity, the dNTP analogue leaving group basicity, and the precise configuration of the C-X atom in R and S stereoisomers on the rate-determining step ( k). The dNTP analogues provide a range of leaving group basicity and steric properties by virtue of monohalogen, dihalogen, or methyl substitution at the carbon atom bridging the β,γ-bisphosphonate that mimics the natural pyrophosphate leaving group in dNTPs.

Base-Mediated Generation of Ketenimines from Ynamides: Direct Access to Azetidinimines by an Imino-Staudinger Synthesis.

Ynamides were used as precursors for the in situ generation of highly reactive ketenimines that could be trapped with imines in a [2+2] cycloaddition. This imino-Staudinger synthesis led to a variety of imino-analogs of β-lactams, namely azetidinimines (20 examples), that could be further functionalized through a broad range of transformations.

At a supra-physiological concentration, human sexual hormones act as quorum-sensing inhibitors.

N-acylhomoserine lactone (AHL)-mediated quorum-sensing (QS) regulates virulence functions in plant and animal pathogens such as Agrobacterium tumefaciens and Pseudomonas aeruginosa. A chemolibrary of more than 3500 compounds was screened using two bacterial AHL-biosensors to identify QS-inhibitors (QSIs). The purity and structure of 15 QSIs selected through this screening were verified using HPLC MS/MS tools and their activity tested on the A.

Copper(I) catalyzed regioselective asymmetric alkoxyamination of aryl enamide derivatives.

The copper(I) catalyzed reaction of an enamide with an iminoiodane, in the presence of an alcohol, triggers the direct alkoxyamination of the double bond. This transformation represents a straightforward access to α-amino aminals in a completely regio- and diastereoselective manner. Use of a chiral Box ligand allows this reaction to be carried out in an enantioselective fashion.