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.

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.