Identification of a mitochondrial target of thiazolidinedione insulin sensitizers (mTOT)--relationship to newly identified mitochondrial pyruvate carrier proteins.

Thiazolidinedione (TZD) insulin sensitizers have the potential to effectively treat a number of human diseases, however the currently available agents have dose-limiting side effects that are mediated via activation of the transcription factor PPARγ. We have recently shown PPARγ-independent actions of TZD insulin sensitizers, but the molecular target of these molecules remained to be identified. Here we use a photo-catalyzable drug analog probe and mass spectrometry-based proteomics to identify a previously uncharacterized mitochondrial complex that specifically recognizes TZDs.

Transcriptional repression and RNA silencing act synergistically to demonstrate the function of the eleventh component of the vaccinia virus entry-fusion complex.

Poxviruses have an elaborate system for infecting cells comprising several proteins for attachment and a larger number dedicated to membrane fusion and entry. Thus far, 11 proteins have been identified as components of the vaccinia virus (VACV) entry-fusion complex (EFC), and 10 of these proteins have been shown to be required for entry. J5, the remaining functionally uncharacterized component of the complex, is conserved in all poxviruses, has a predicted C-terminal transmembrane domain, and is an N-terminally truncated paralog of two other EFC proteins.

Interaction between the G3 and L5 proteins of the vaccinia virus entry-fusion complex.

The vaccinia virus entry-fusion complex (EFC) consists of 10 to 12 proteins that are embedded in the viral membrane and individually required for fusion with the cell and entry of the core into the cytoplasm. The architecture of the EFC is unknown except for information regarding two pair-wise interactions: A28 with H2 and A16 with G9. Here we used a technique to destabilize the EFC by repressing the expression of individual components and identified a third pair-wise interaction: G3 with L5.

Activity of recombinant cysteine-rich domain proteins derived from the membrane-bound MUC17/Muc3 family mucins.

Background: The membrane-bound mucins, MUC17 (human) and Muc3 (mouse), are highly expressed on the apical surface of intestinal epithelia and have cytoprotective properties. Their extracellular regions contain two EGF-like Cys-rich domains (CRD1 and CRD2) connected by an intervening linker segment with SEA module (L), and may function to stimulate intestinal cell restitution. The purpose of this study was to determine the effect of size, recombinant host source, and external tags on mucin CRD1-L-CRD2 protein activity.

Three-dimensional reconstruction of the valyl-tRNA synthetase/elongation factor-1H complex and localization of the delta subunit.

Eukaryotic valyl-tRNA synthetase (ValRS) and the heavy form of elongation factor 1 (EF-1H) are isolated as a stable high molecular mass complex that catalyzes consecutive steps in protein biosynthesis--aminoacylation of tRNA and its transfer to elongation factor. Herein is the first three-dimensional structure of the particle as calculated from electron microscopic images of negatively stained samples of the human ValRS/EF-1H complex. The ca.

A three-dimensional working model of the multienzyme complex of aminoacyl-tRNA synthetases based on electron microscopic placements of tRNA and proteins.

It has become evident that the process of protein synthesis is performed by many cellular polypeptides acting in concert within the structural confines of protein complexes. In multicellular eukaryotes, one of these assemblies is a multienzyme complex composed of eight proteins that have aminoacyl-tRNA synthetase activities as well as three non-synthetase proteins (p43, p38, and p18) with diverse functions. This study uses electron microscopy and three-dimensional reconstruction to explore the arrangement of proteins and tRNA substrates within this "core" multisynthetase complex.

Immobilization of a novel antibacterial agent on solid phase and subsequent isolation of EF-Tu.

Screening of our compound collection identified PNU-92560, a 2-[1,3,4]thiadiazolo[3,2-a]pyrimidine-6-carboxamide, as a novel antibacterial agent. Extensive analogue development identified that the 2-position of the thiadiazole could be functionalized with a linker that would allow the compound to be attached to a solid support. The extreme insolubility of the analogues prevented the mechanism of action for these compounds to be determined utilizing traditional methodology.

Isolation and characterization of human nuclear and cytosolic multisynthetase complexes and the intracellular distribution of p43/EMAPII.

In this study, the human multienzyme aminoacyl-tRNA synthetase "core" complex has been isolated from the nuclear and cytosolic compartments of human cells and purified to near homogeneity. It is clear from the polypeptide compositions, stoichiometries, and three-dimensional structures that the cytosolic and nuclear particles are very similar to each other and to the particle obtained from rabbit reticulocytes. The most significant difference observed via aminoacylation activity assays and densitometric analysis of electrophoretic band patterns is a lower amount of glutaminyl-tRNA synthetase in the human particles.

Co-crystallization of Staphylococcus aureus peptide deformylase (PDF) with potent inhibitors.

In bacteria the biosynthesis of all nascent polypeptides begins with N-formylmethionine. The post-translational removal of the N-formyl group is carried out by peptide deformylase (PDF). Processing of the N-formyl group from critical bacterial proteins is required for cell survival.

Crystal structure of type II peptide deformylase from Staphylococcus aureus.

The first crystal structure of Class II peptide deformylase has been determined. The enzyme from Staphylococcus aureus has been overexpressed and purified in Escherichia coli and the structure determined by x-ray crystallography to 1.9 A resolution.