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Recent Research Papers of Note: Mar 19, 2009


Journal: Electrophoresis, March 16 [Epub ahead of print]
Title: Proteomic analysis for process development and control of therapeutic protein separation from human plasma
Authors: X Yang; J Clifton; F Huang; S Kovac; DC Hixson; D Josic

Authors demonstrate a new two-step process for plasma separation. The two most abundant proteins, HSA and IgG, were depleted in a first step of anion-exchange chromatography using a gel with high capacity. Two fractions containing medium- and low-abundance proteins were re-chromatographed on a smaller column with the same type of gel. Collected fractions were separated by SDS-PAGE and 2D electrophoresis. Excised proteins were tryptically digested and identified by LC-ESI-MS/MS.

Journal: Analytical Chemistry, March 15
Title: Metal-coded affinity tag labeling: A demonstration of analytical robustness and suitability for biological applications
Authors: R Ahrends; S Pieper; B Neumann; C Scheler; MW Linscheid

According to the abstract, metal-coded affinity tags, or MeCAT, were developed to provide a "robust tool for the quantification of peptides and proteins by utilizing lanthanide-harboring metal tags." The study presented here investigates the compatibility of MeCAT labeling to workflows such as nano-LC-ESI MS. They especially focused on the separation behavior of labeled peptides and the dynamic range of detection and peptide charge distribution. The stability of MeCAT under harsh analytical conditions was also investigated.

Journal: Electrophoresis, March 12 [Epub ahead of print]
Title: Comparison of 1D and 2D LC MS-MS methods for proteomic analysis of human serum
Authors: M Gilar; P Olivova; AB Chakraborty; A Jaworski; SJ Geromanos; JC Gebler

Authors used 1D and 2D methods for proteome analysis of undepleted human serum. Applying selected data filtration, they identified 52 proteins based on 316 peptides in serum in a 1D LC setup, 184 proteins out of 1,036 peptides in an RP-RP 2D LCsetup, and 142 proteins out of 905 peptides in an SCX-RP 2D LC setup. The authors also compared the 2D LC methods for proteomic analysis.

Journal: Analytical Chemistry, March 10 [Epub ahead of print]
Title: Application of a combined weak cation-exchange/crown ether columns: First demonstrations of a versatile tool for proteome subselection
Authors: R Tuytten; B Ruttens; K Gheysen; K Sandra; K de Cremer; D Vlieghe; N van Landuyt; G Thomas; JC Martins; P Sandra; K Kas; K Verleysen

The paper introduces their column for proteomics use. According to the abstract, the 18-crown-6 ether functionality is "well-known" to selectively complex ammonium and monoalkylammonium ions, "which should make this column highly suitable to trap peptides with free alpha-NH2 or free epsilon- NH2 groups from the lysine side chains." The authors tested the mechanism in an N-teromics setup for the enrichment of deliberately acetylated protein N-terminal peptides from a serum digest and demonstrated that peptides with free alpha-NH2 groups and peptides with alpha-amino-acetylated groups can be separated from each other using their column.

Journal: Journal of Proteome Research, March 6
Title: Human serum proteins fractionated by preparative partition chromatography prior to LC-ESI-MS/MS
Authors: M Tucholska; P Bowden; K Jacks; P Zhu; S Furesz; M Dumbrovsky; J Marshall

Authors describe a method to get at low-abundance proteins and show that their approach resulted in high-stringency tryptic peptides identified by LC-ESI-MS/MS. Based on their results, they said that "the depletion of albumin or IgG was not necessary prior to LC-MS/MS and that multiple forms of protein chromatography will be useful for complete identification of blood proteins."

Journal: Journal of Proteome Research, March 3 [Epub ahead of print]
Title: Post analysis data acquisition for the interative MS/MS sampling of proteomics mixtures
Authors: MR Hoopmann; GE Merrihew; PD von Haller; MJ MacCoss

Despite the routine identification of peptides by microcapillary LC-MS/MS, many peptides within the detection limit of the mass spectrometer remain unidentified due to limitations in MS/MS sample speed. Authors have developed an automated approach using the mass spectra from high-resolution muLC-MS data to define "the molecular species present in the mixture and directs the acquisition of MS/MS spectra to precursors that were missed in prior analyses."

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Journal: Molecular & Cellular Proteomics, March 2 [Epub ahead of print]
Title: Spectral profiles: A novel representation of tandem mass spectra and its applications for de novo peptide sequencing and identification
Authors: S Kim; N Bandeira; PA Pevzner

Authors say that despite the many efforts during the past decade, "the progress in de novo peptide sequencing has been slow with only 30-45 percent of all peptides being correctly constructed." Accurate full-length peptide sequencing may be unattainable for some spectra, they said, and demonstrate how to accurately sequence gapped peptides instead, which they said "are nearly as useful as full-length peptides for error-tolerant database searches." Their MS-Profile tool uses spectral profiles, a new representation of MS/MS spectra, to generate gapped peptides "that are longer and more accurate than peptide sequence tags of length 3 traditionally used to speed up database searches in proteomics."

Journal: The Journal of Pathology, March
Title: Development and validation of a novel protein extraction methodology for quantitation of protein expression in formalin-fixed paraffin-embedded tissues using Western blotting
Authors: NJ Nirmalan; P Harnden; PJ Selby; RE Banks

Presented is a heat-induced antigen retrieval strategy using SDA-containing Laemmli buffer for efficient protein recovery from formalin-fixed tissues for analysis by Western blotting. The paper also presents protocol optimization and comparison of extraction efficacies with frozen tissues and current methodology.

Journal: Proteomics, March
Title: Improving sensitivity in proteome studies by analysis of false discovery rates for multiple search engines
Authors: AR Jones; JA Siepen; SJ Hubbard; NW Paton

Authors have developed a search engine-independent score, based on false discovery rates, which allows peptide identifications from different search engines to be combined. They call the score FDR Score. "The results demonstrate that the observed FDR is significantly different when analyzing the set of identifications made by all three search engines, by each pair of search engines or by a single search engine," according to the abstract.

Journal: Proteomics, March
Title: POSTMan (Post-translational modification analysis), a software application for PTM discovery
Authors: MØ Arntzen; CL Osland; CR Raa; R Kopperud; SO Døskeland; AE Lewis; CS D'Santos

According to the authors, post-translationally modified peptides present in low concentrations are often not selected for collision-induced dissociation resulting in no sequence information for these peptides. They have developed software, POSTMan, which allows post-translationally modified peptides to be targeted for fragmentation. The software aligns LC-MS runs between individual runs or within a single run and isolates pairs of peptides that differ by a user-defined mass difference.

Journal: Rapid Communications in Mass Spectrometry: RCM, March
Title: Top-down proteomics with a quadrupole time-of-flight mass spectrometer and collision-induced dissociation
Authors: A Armirotti; U Benatti; G Damonte

Authors demonstrate that with slight modifications to the instrumental parameters, CID can be performed on a quadrupole time-of-flight MS not originally designed for such a purpose. Protein identification is done with both N- and C-terminal sequence tags and Blast database searches. The accurate mass measurement of multiply charged fragment ions supplements the limited set of cleavage sites and provides a "high degree" of sequence coverage, they said, and PTM issues can be addressed.

Journal: Proteomics, Feb. 20 [Epub ahead of print]
Title: Strategy for surveying the proteome using affinity proteomics and mass spectrometry
Authors: C Wingren; P James; CA Borrebaeck

While antibody-based microarrays are a "rapidly evolving technology" the number of antibodies included on the microarrays is a "key limiting factor," the authors said in the abstract. To overcome this "and to be able to perform in-depth global proteome surveys" they propose interface affinity proteomics with MS-based readout. In their approach, they have defined a range of peptide motifs, each being present in five to 100 different proteins. "In this manner, 100 antibodies, binding 100 different motifs commonly distributed among different proteins, would potentially target a protein cluster of [10,000] individual molecules" or around half of the non-redundant human proteome, according to the abstract.

The Scan

Study Examines Insights Gained by Adjunct Trio RNA Sequencing in Complex Pediatric Disease Cases

Researchers in AJHG explore the diagnostic utility of adding parent-child RNA-seq to genome sequencing in dozens of families with complex, undiagnosed genetic disease.

Clinical Genomic Lab Survey Looks at Workforce Needs

Investigators use a survey approach in Genetics in Medicine Open to assess technologist applications, retention, and workforce gaps at molecular genetics and clinical cytogenetics labs in the US.

Study Considers Gene Regulatory Features Available by Sequence-Based Modeling

Investigators in Genome Biology set sequence-based models against observational and perturbation assay data, finding distal enhancer models lag behind promoter predictions.

Genetic Testing Approach Explores Origins of Blastocyst Aneuploidy

Investigators in AJHG distinguish between aneuploidy events related to meiotic missegregation in haploid cells and those involving post-zygotic mitotic errors and mosaicism.