Discovery of a novel class of boron-based antibacterials with activity against gram-negative bacteria.

Gram-negative bacteria cause approximately 70% of the infections in intensive care units. A growing number of bacterial isolates responsible for these infections are resistant to currently available antibiotics and to many in development. Most agents under development are modifications of existing drug classes, which only partially overcome existing resistance mechanisms.

A nonradioactive high-throughput assay for screening and characterization of adenylation domains for nonribosomal peptide combinatorial biosynthesis.

Adenylation domains are critical enzymes that dictate the identity of the amino acid building blocks to be incorporated during nonribosomal peptide (NRP) biosynthesis. NRPs display a wide range of biological activities and are some of the most important drugs currently used in clinics. Traditionally, activity of adenylation domains has been measured by radioactive ATP-[32P]pyrophosphate (PP(i)) exchange assays.

Characterization of TioF, a tryptophan 2,3-dioxygenase involved in 3-hydroxyquinaldic acid formation during thiocoraline biosynthesis.

Thiocoraline is a thiodepsipeptide antitumor agent that belongs to the family of bisintercalator natural products that bind duplex DNA through their two planar intercalating moieties. In thiocoraline, the 3-hydroxyquinaldic acid (3HQA) chromophores required for intercalation are derived from L-Trp. We have expressed the Micromonospora sp.

Activation of D-tyrosine by Bacillus stearothermophilus tyrosyl-tRNA synthetase: 1. Pre-steady-state kinetic analysis reveals the mechanistic basis for the recognition of D-tyrosine.

Tyrosyl-tRNA synthetase (TyrRS) is able to catalyze the transfer of both l- and d-tyrosine to the 3' end of tRNA(Tyr). Activation of either stereoisomer by ATP results in formation of an enzyme-bound tyrosyl-adenylate intermediate and is accompanied by a blue shift in the intrinsic fluorescence of the protein. Single turnover kinetics for the aminoacylation of tRNA(Tyr) by D-tyrosine were monitored using stopped-flow fluorescence spectroscopy.

Activation of D-tyrosine by Bacillus stearothermophilus tyrosyl-tRNA synthetase: 2. Cooperative binding of ATP is limited to the initial turnover of the enzyme.

The activation of D-tyrosine by tyrosyl-tRNA synthetase has been investigated using single and multiple turnover kinetic methods. In the presence of saturating concentrations of D-tyrosine, the activation reaction displays sigmoidal kinetics with respect to ATP concentration under single turnover conditions. In contrast, when the kinetics for the activation reaction are monitored using a steady-state (multiple turnover) pyrophosphate exchange assay, Michaelis-Menten kinetics are observed.

Purification strategies for microbial lipases.

Microbial lipases today occupy a place of prominence among biocatalysts owing to their ability to catalyze a wide variety of reactions in aqueous and non-aqueous media. The chemo-, regio- and enantio-specific behaviour of these enzymes has caused tremendous interest among scientists and industrialists. Lipases from a large number of bacterial, fungal and a few plant and animal sources have been purified to homogeneity.