Application of an allosteric model to describe the interactions among retinol binding protein 4, transthyretin, and small molecule retinol binding protein 4 ligands.

Retinol binding protein 4 (RBP4) is a serum protein that serves as the major transport protein for retinol (vitamin A). Recent reports suggest that elevated levels of RBP4 are associated with insulin resistance and that insulin sensitivity may be improved by reducing serum RBP4 levels. This can be accomplished by administration of small molecules, such as fenretinide, that compete with retinol for binding to RBP4 and disrupt the protein-protein interaction between RBP4 and transthyretin (TTR), another serum protein that protects RBP4 from renal clearance.

A high-throughput microfluidic assay for SH2 domain-containing inositol 5-phosphatase 2.

SH2 domain-containing inositol 5-phosphatase 2 (SHIP2) is a potential drug target for the treatment of type 2 diabetes. This enzyme serves as a negative regulator of insulin-mediated signal transduction by catalyzing the dephosphorylation of the second messenger lipid molecule phosphatidylinositol 3,4,5-triphosphate. Traditionally, assays for phosphoinositide phosphatases such as SHIP2 have relied on radiolabeled phosphatidylinositol-containing lipid membranes and chromatographic separation of labeled phospholipid substrate from product by thin-layer chromatography.

Use of a modified tryptophanase promoter to direct high-level expression of foreign proteins in E. coli.

We have modified the tryptophanase promoter (PtnaA) for use as a temperature-independent promoter for the production of recombinant proteins. Although any protein will have a temperature range in which its expression is optimal, we find the tryptophanase promoter functions at all physiologically relevant temperatures (20 degrees C to 42 degrees C). Induction at temperatures below 37 degrees C avoids eliciting the heat-shock response and may favor the production of protein in the soluble state.

Design and evaluation of a two-stage, cyclic, recombinant fermentation process.

A two-stage, cyclic fed-batch fermentation process to produce recombinant human lymphokine was designed. The organism used in the study was Escherichia coli K-12 containing a temperature-sensitive walkaway plasmid bearing an insert which codes for a human lymphokine. Transcription of the recombinant gene is controlled by a lambda repressor/pL promoter system.