Role of pro-oncogenic protein disulfide isomerase (PDI) family member anterior gradient 2 (AGR2) in the control of endoplasmic reticulum homeostasis.

The protein-disulfide isomerase (PDI) family member anterior gradient 2 (AGR2) is reportedly overexpressed in numerous cancers and plays a role in cancer development. However, to date the molecular functions of AGR2 remain to be characterized. Herein we have identified AGR2 as bound to newly synthesized cargo proteins using a proteomics analysis of endoplasmic reticulum (ER) membrane-bound ribosomes.

Proteomic analysis of the transitional endoplasmic reticulum in hepatocellular carcinoma: an organelle perspective on cancer.

The transitional endoplasmic reticulum (tER) is composed of both rough and smooth ER membranes and thus participates in functions attributed to both these two subcellular compartments. In this paper we have compared the protein composition of tER isolated from dissected liver tumor nodules of aflatoxin B1-treated rats with that of tER from control liver. Tandem mass spectrometry (MS), peptide counts and immunoblot validation were used to identify and determine the relative expression level of proteins.

Proteomics analysis of liver pathological calcification suggests a role for the IQ motif containing GTPase activating protein 1 in myofibroblast function.

To date the cellular and molecular mechanisms by which liver pathological calcifications occur and are regulated are poorly investigated. To study the mechanisms linked to their appearance, we performed a proteomics analysis of calcified liver samples. To this end, human liver biopsies collected in noncalcified (N), precalcified (P), and calcified (C) areas of the liver were subjected to weak ion exchange chromatography, SDS-PAGE, and LC-ESI MS/MS analyses.

A Bayesian approach to peptide identification using accurate mass and time tags from LC-FTICR-MS proteomics experiments.

In high-throughput proteomics, one promising approach presently being explored is the Accurate Mass and Time (AMT) tag approach, in which reversed-phase liquid chromatography coupled to high accuracy mass spectrometry provide measurements of both the masses and chromatographic retention times of tryptic peptides in complex mixtures. These measurements are matched to the mass and predicted retention times of peptides in library. There are two varieties of peptides in the library: peptides whose retention time predictions are derived from previous peptide identifications and therefore are of high precision, and peptides whose retention time predictions are derived from a sequence-based model and therefore have lower precision.

Room temperature stability of injectable succinylcholine dichloride.

The purpose of this study was to determine the room temperature stability over a period of several months of commercially available intravenous succinylcholine dichloride (Quelicin, 20 mg/mL) in vials. A previously validated electro-spray tandem mass spectrometry method developed for the determination of succinylcholine dichloride in plasma was used. This method was based upon a stable isotope dilution assay using hexadeuterosuccinylcholine diiodide as the internal standard and was shown to be specific, sensitive, and reproducible.

Quantification of uncertainty of peptide retention time predictions from a sequence-based model in LC-MS/MS proteomics experiments.

In high-throughput mass spectrometry-based proteomics, it is necessary to employ separations to reduce sample complexity prior to mass spectrometric peptide identification. Interest has begun to focus on using information from separations to aid in peptide identification. One of the most common separations is reversed-phase liquid chromatography, in which peptides are separated on the basis of their chromatographic retention time.

Proteomic analysis of tyrosine phosphorylation during human liver transplantation.

Background: Ischemia-reperfusion (I/R) causes a dramatic reprogramming of cell metabolism during liver transplantation and can be linked to an alteration of the phosphorylation level of several cellular proteins. Over the past two decades, it became clear that tyrosine phosphorylation plays a pivotal role in a variety of important signalling pathways and was linked to a wide spectrum of diseases. Functional profiling of the tyrosine phosphoproteome during liver transplantation is therefore of great biological significance and is likely to lead to the identification of novel targets for drug discovery and provide a basis for novel therapeutic strategies.

Proteomic analysis of ischemia-reperfusion injury upon human liver transplantation reveals the protective role of IQGAP1.

Ischemia-reperfusion injury (IRI) represents a major determinant of liver transplantation. IRI-induced graft dysfunction is related to biliary damage, partly due to a loss of bile canaliculi (BC) integrity associated with a dramatic remodeling of actin cytoskeleton. However, the molecular mechanisms associated with these events remain poorly characterized.

Characterization of an RNA granule from developing brain.

In brain, mRNAs are transported from the cell body to the processes, allowing for local protein translation at sites distant from the nucleus. Using subcellular fractionation, we isolated a fraction from rat embryonic day 18 brains enriched for structures that resemble amorphous collections of ribosomes. This fraction was enriched for the mRNA encoding beta-actin, an mRNA that is transported in dendrites and axons of developing neurons.

Multicomponent internal recalibration of an LC-FTICR-MS analysis employing a partially characterized complex peptide mixture: systematic and random errors.

In high-throughput proteomics, a promising current approach is the use of liquid chromatography coupled to Fourier transform ion cyclotron resonance mass spectrometry (LC-FTICR-MS) of tryptic peptides from complex mixtures of proteins. To apply this method, it is necessary to account for any systematic measurement error, and it is useful to have an estimate of the random error expected in the measured masses. Here, we analyze by LC-FTICR-MS a complex mixture of peptides derived from a sample previously characterized by LC-QTOF-MS.

Increasing peptide identification in tandem mass spectrometry through automatic function switching optimization.

Comprehensive proteomic studies that employ MS directed peptide sequencing are limited by optimal peptide separation and MS and tandem MS data acquisition routines. To identify the optimal parameters for data acquisition, we developed a system that models the automatic function switching behavior of a mass spectrometer using an MS-only dataset. Simulations were conducted to characterize the number and the quality of simulated fragmentation as a function of the data acquisition routines and used to construct operating curves defining tandem mass spectra quality and the number of peptides fragmented.

Tandem MS analysis of brain clathrin-coated vesicles reveals their critical involvement in synaptic vesicle recycling.

Tandem MS has identified 209 proteins of clathrin-coated vesicles (CCVs) isolated from rat brain. An overwhelming abundance of peptides were assigned to the clathrin coat with a 1:1 stoichiometry observed for clathrin heavy and light chains and a 2:1 stoichiometry of clathrin heavy chain with clathrin adaptor protein heterotetramers. Thirty-two proteins representing many of the known components of synaptic vesicles (SVs) were identified, supporting that a main function for brain CCVs is to recapture SVs after exocytosis.

The model B6(dom1) minor histocompatibility antigen is encoded by a mouse homolog of the yeast STT3 gene.

The B6(dom1) minor histocompatibility antigen (MiHA) is a model antigen, since it is both the epitome of an immunodominant epitope and an ideal target for adoptive cancer immunotherapy. Based on DNA sequencing and MS/MS analyses, we report that B6(dom1) corresponds to amino acids 770-778 (KAPDNRETL) of a protein we propose to call SIMP (source of immunodominant MHC-associated peptides) that is encoded by a mouse homolog of the yeast STT3gene. STT3, a member of the oligosaccharyltransferase complex, is essential for cell proliferation.

Concentration-effect relation of succinylcholine chloride during propofol anesthesia.

Background: The pharmacokinetics and pharmacodynamics of succinylcholine were studied simultaneously in anesthetized patients to understand why the drug has a rapid onset and short duration of action. A quantitative model describing the concentration-effect relation of succinylcholine was proposed. The correlation between hydrolysis in plasma and elimination was also examined.

Enthoprotin: a novel clathrin-associated protein identified through subcellular proteomics.

Despite numerous advances in the identification of the molecular machinery for clathrin-mediated budding at the plasma membrane, the mechanistic details of this process remain incomplete. Moreover, relatively little is known regarding the regulation of clathrin-mediated budding at other membrane systems. To address these issues, we have utilized the powerful new approach of subcellular proteomics to identify novel proteins present on highly enriched clathrin-coated vesicles (CCVs).