Intracellular Binding of Terfenadine Competes with Its Access to Pancreatic ß-cell ATP-Sensitive K Channels and Human ether-à-go-go-Related Gene Channels.

Most blockers of both hERG (human ether-à-go-go-related gene) channels and pancreatic ß-cell ATP-sensitive K (K) channels access their binding sites from the cytoplasmic side of the plasma membrane. It is unknown whether binding to intracellular components competes with binding of these substances to K channels. The whole-cell configuration of the patch-clamp technique, a laser-scanning confocal microscope, and fluorescence correlation spectroscopy (FCS) were used to study hERG channels expressed in HEK (human embryonic kidney) 293 cells and K channels from the clonal insulinoma cell line RINm5F.

An irreversible and kinetically controlled process: thermal induced denaturation of L-2-hydroxyisocaproate dehydrogenase from Lactobacillus confusus.

The thermal denaturation of Lactobacillus confusus L-2-Hydroxyisocaproate Dehydrogenase (L-HicDH) has been studied by Differential Scanning Calorimetry (DSC). The stability of this enzyme has been investigated at different pH conditions. The results of this study indicate that the thermal denaturation of this enzyme is irreversible and the T(m) is dependent on the scan-rate, which suggests that the denaturation process of L-HicDH is kinetically determined.

Cloning, expression, purification and preliminary crystallographic characterization of a shikimate dehydrogenase from Corynebacterium glutamicum.

The shikimate dehydrogenase from Corynebacterium glutamicum has been cloned into an Escherichia coli expression vector, overexpressed and purified. Native crystals were obtained by the vapour-diffusion technique using 2-methyl-2,4-pentanediol as a precipitant. The crystals belong to the centred monoclinic space group C2, with unit-cell parameters a = 118.

High level expression and single-step purification of hexahistidine-tagged L-2-hydroxyisocaproate dehydrogenase making use of a versatile expression vector set.

Affinity tags as fusions to the N- or C-terminal part of proteins are valuable tools to facilitate the production and purification of proteins. In many cases, there may be the necessity to remove the tag after protein preparation to regain activity. Removal of the tag is accomplished by insertion of a unique amino acid sequence that is recognized and cleaved by a site specific protease.