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Recent Research Papers of Note in Proteomics: Sep 18, 2008

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Journal: Journal of Proteome Research, Sept. 13 [Epub ahead of print]
Title: Adaptive discriminant function analysis and reranking of MS/MS database search results for improved peptide identification in shotgun proteomics.
Authors: Y Ding; H Choi; AI Nesvizhskii
 
Authors investigate the limits of PeptideProphet, a tool commonly used for peptide identification, and describe an adaptive method in which a new discriminant function is “learned from the data in an iterative fashion” to address the limitations, according to the abstract. The authors extend the modeling framework to “go beyond the top scoring peptide assignment per spectrum,” they said, and investigate the effect of clustering the spectra based on their spectrum quality score followed by cluster-specific mixture modeling.
 

 
Journal: Analytical Chemistry, Sept. 13 [Epub ahead of print]
Title: Sequential interval motif search: Unrestricted database surveys of global MS/MS data sets for detection of putative post-translational modifications.
Authors: J Liu; A Erassov; P Halina; M Canete; ND Vo; C Chung; G Cagney; A Ignatchenko; V Fong; A Emili
 
The article describes a new algorithm, Sequential Motif Interval Search, or SIMS, for the detection of unanticipated putative post-translational modification. SIMS interprets pairs of product ion peaks “representing potential amino acid residues, or ‘intervals,’ as a means of mapping PTMs or substitutions in a blind database search mode,” according to the abstract.
 
Authors also developed an effective heuristic software program to evaluate, rank, and filter optimal combinations of relevant intervals to detect candidate sequences and any associated PTMs or polymorphisms from MS/MS spectra. SIMS was benchmarked against annotated reference spectral data sets “and compared favorably with, and was complementary to current state-of-the-art methods,” the authors said. 
 

 
Journal: Proteome Science, Sept. 12 [Epub ahead of print]
Title: A simple and reliable protocol for mouse serum proteome profiling studies by use of two-dimensional electrophoresis and MALDI TOF/TOF mass spectrometry
Authors: MS Ritorto, J Borlak
 
Addressing the need for better protocols for the identification of serum proteins, authors report a combination of two proteomic strategies, acidic and neutral part of 2D gels, and an application of two optimized matrix preparations for MALDI-MS to simplify serum proteome mapping. Authors report their protocol allowed automated data acquisition for both alpha-ciano-4-hydroxycinnamic acid matrix deposition and dihydroxybenzoic acid and simplified MS-data acquisition.
 

 
Journal: Journal of Proteome Research, Sept. 12 [Epub ahead of print]
Title: Improved sequence tag generation method for peptide identification in tandem mass spectrometry.
Authors: X Cao; AI Nesvizhskii
 
While sequence tag-based peptide identification methods are a “promising” alternative to traditional database search approaches, the authors said that a more comprehensive analysis, optimization, and comparison with established methods are needed for widespread adoption of tag-based methods. Using the InsPecT open-source code base, they present an “improved sequence tag generation method that directly incorporates multi-charged fragment ion peaks present in many tandem mass spectra of higher charge states,” according to the abstract. They further investigate the performance of sequence tagging under different settings using control datasets from five different types of mass spectrometers and a complex phosphopeptide-enriched sample.
 

 
Journal: Analytical Chemistry, Sept. 11 [Epub ahead of print]
Title: De novo sequencing of unique sequence tags for discovery of post-translational modifications of proteins.
Authors: Y Shen; N Toliæ; KK Hixson; SO Purvine; GA Anderson; RD Smith
 
Described is a de novo sequencing approach for protein modifications based on identification of the proteome UStags. The de novo information was obtained from Fourier transform tandem mass spectrometry data for peptides and polypeptides from a yeast lysate. The DNA-predicted database protein sequences were compared to the UStags and the differences observed across or in the UStags were used to infer possible sequence modifications. The approach allowed the authors to uncover unexpected variances within several yeast protein sequences due to amino acid mutations and/or multiple modifications to the predicted protein sequences.
 

 
Journal: Journal of Proteome Research, Sept. 11 [Epub ahead of print]
Title: pICarver: A software tool and strategy for peptides isoelectric focusing
Authors: AR Vaezzadeh; C Hernandez; O Vadas; JJ Deshusses; P Lescuyer; F Lisacek; DF Hochstrasser
 
According to the abstract, the use of isoelectric focusing as the first dimension of separation is a new trend in shotgun proteomics. The authors present the development of a new tool and strategy “that generates a fractionation scheme resulting in almost even distribution of peptides per fraction,” according to the abstract. The pICarver software also increases the throughput of the approach by reducing the number of fractions and merging the peptide-poor regions.
 

 
Journal: Proteomics, Sept. 9 [Epub ahead of print]
Title: Improvements of TArgeted multiplex mass spectrometry IMaging
Authors: G Thiery; E Anselmi; A Audebourg; E Darii; M Abarbri; B Terris; JC Tabet; IG Gut
 
Authors report an improvement of Targeted multiplex MS Imaging, or TAMSIM, technology, by attaching photocleavable mass tags to antibodies. Histological sections are stained analogous to standard immunohistochemical procedures with chemiluminescent or fluorescent detection “with the sole difference that multiple antibodies each with a distinct mass tag are used in a single reaction,” according to the abstract. Laser pulse at 355 nm without added matrix is used to release mass tags from their respective antibodies, and after scanning MS images are created for each tag mass.
 

 
Journal: Molecular Cell, Sept. 5
Title: Combined use of RNAi and quantitative proteomics to study gene function in Drosophila
Authors: T Bonaldi; T Straub; J Cox; C Kumar; PB Becker; M Mann
 
Authors applied stable isotope labeling by amino acids in cell culture, or SILAC, and RNAi to Drosophila SL2 cells. After knockdown of ISWI, an ATP-hydrolyzing motor of different chromatin remodeling complexes, they obtained a quantitative proteome of more than 4,000 proteins. ISWI itself was reduced 10-fold as quantified by SILAC, and several hundred proteins were regulated and clustered into distinct functional categories.
 

 
Journal: Journal of Proteome Research, Sept. [Epub ahead of print]
Title: Fractionation of complex protein mixture by virtual three-dimensional liquid chromatography based on combined pH and salt steps
Authors: ZB Ning; QR Li; J Dai; RX Li; CH Shieh; R Zeng
 
Authors describe a chromatographic method for sample fractionation called virtual three-dimensional chromatography. They alternated elution with different pHs and salt concentrations, implementing pH and salt steps by turns on a single strong cation exchange column. They further fractionated and desalted proteins with a reversed-phase column tandem connected behind. They report 1,933 protein groups covering a wide dynamic range resulting from a single experiment with their approach.
 

 
Journal: Biotechniques, September
Title: Identification of highly expressed, soluble proteins using an improved, high-throughput pooled ORF expression technology.
Authors: T Waybright; W Gillette; D Esposito; R Stephens; D Lucas; J Hartley; T Veenstra
 
Described is an improved pooled open-reading frame expression technology using recombinatorial cloning and solution-based tandem mass spectrometry to identify ORFS yielding high levels of soluble, purified proteins when expressed in E. coli. The authors used their method to subclone, purify, and transfect three identical pools of 512 human ORFs into three separate E. coli cultures. After bulk expression and purification, the proteins from each pool were digested into tryptic peptides, and each sample was analyzed in triplicate using reversed-phase HPLC, coupled directly online with MS/MS.
 
By calculating the average exponentially modified protein abundance index of each protein across the three protein pools, they determined the abundance of each protein. Human protein exhibiting high emPAI values were subjected to small-scale expression and purification. These clones showed high levels of expression of soluble protein, and proteins that were not observed by LC-MS/MS did not show any detectable expression in small-scale validation studies.
 

 
Journal: Journal of Proteome Research, September
Title: Robust prediction of the MASCOT score for an improved quality assessment in mass spectrometric proteomics
Authors: T Koenig; BH Menze; M Kirchner; F Monigatti; K C Parker; T Patterson; JJ Steen; FA Hamprecht; H Steen
 
Described is a continuous quality score that can be “computed very quickly and that can be considered an approximation of the MASCOT score in case of a correct identification,” according to the abstract. The score can be used to reject low-quality spectra and can be calibrated automatically on-site without a manually generated training set.
 

 
Journal: Journal of Proteome Research, September
Title: Quantitative analysis of redox-sensitive proteome with DIGE and ICAT
Authors: C Fu; J Hu; T Liu: T Ago; J Sadoshima; H Li
 
Authors set out to compare difference gel electrophoresis to isotope-coded affinity tag for the discovery of redox-sensitive proteins in heat tissues. They found the two methods to be largely complementary: For DIGE analysis, both NF-kappaB-repressing protein and epoxide hydrolase were sensitive to H2O2 oxidation, while with ICAT, specific cysteines within sacroplasmic endoplasmic reticulum calcium ATPase 2 and voltage-dependent anion-selective channel protein were sensitive to H2O2 oxidation.
 
“From these analyses, we conclude that both methods should be employed for proteome-wide studies, to maximize the possibility of identifying proteins containing redox-sensitive cysteinyl thiols in complex biological systems,” the authors said in the abstract.
 

 
Journal: Journal of Proteome Research, September
Title: Immobilized metal affinity chromatography revisited: pH/acid control toward high selectivity in phosphoproteomics
Authors: CF Tsai; YT Wang; YR Chen; CY Lai; PY Lin; KT Pan; JY Chen; KH Khoo; YJ Chen
 
Authors report a simple pH/acid control method “that addresses the poor specificity seriously criticized in IMAC,” according to the abstract. By controlling salt, pH, and the structure and concentration of organic acid, a revised one-step IMAC method with low sample loss can be designed. The protocols can be adapted to other fully automated or manual solid supports for large-scale identification of the phosphoproteome, according to the authors.
 

 
Journal: Journal of Proteome Research, September
Title: Precursor-ion mass re-estimation improves peptide identification on hybrid instruments
Authors: R Luethy; DE Kessner; JE Katz; B Maclean; R Grothe; K Kani; V Faca; S Pitteri; S Hanash; DB Agus; P Mallick
 
While hybrid mass spectrometers have become important research tools due to their ability to identify larger number of peptides with higher confidence than competing instruments, the authors said that they have noted parent masses deviate 171 parts per mole, on average, for ion-trap data directed identifications, and 8 ppm, on average, for preview Fourier transform data directed identifications.
 
To improve these deviations, a program, msPrefix, is introduced “to re-estimate a peptide's parent mass from an associated high-accuracy full-scan survey spectrum,” according to the abstract. In an 18-protein mixture, msPrefix parent mass estimates deviate only 1 ppm, on average, from the identified peptides. In complex mixture experiments, the authors show that more than half of triggered MS/MS spectra “may have had multiple precursor ions and note that spectra with multiple candidate ions are less likely to result in an identification using TANDEM,” according to the abstract. The results demonstrate that integrating msPrefix into traditional shotgun proteomics workflows “significantly improves identification results.”
 

 
Journal: Journal of Proteome Research, September
Title: Rapid sample processing for LC-MS-based quantitative proteomics using high intensity focused ultrasound
Authors: D Lopez-Ferrer; TH Heibeck; K Petritis; KK Hixson; W Qian; ME Monroe; A Mayampurath; RJ Moore; ME Belov; DG Camp; RD Smith
 
Described is a new sample processing workflow for quantitative proteomics applications, using high intensity focused ultrasound to rapidly reduce alkylate cysteines, digest proteins, and label peptides with 18O. Using the workflow, the authors denatured, alkylated, in-solution digested and 18O-labeled mouse plasma proteins in less than 10 minutes for subsequent analysis liquid chromatography-electrospray ionization high resolution mass spectrometry.
 

 
Journal: Proteomics, September
Title: 2-D DIGE and MS/MS analysis of protein serum expression in rats housed in concrete and clay cages in winter.
Authors: JC Kim; JY Kim; SR Yeom; BY Jeon; HZ Hwang; KJ Park; SW Lee
 
In a previous study, the authors looked at the physiological responses of male Sprague-Dawley rats over a four-week exposure to concrete and clay cages and detected no general toxicological changes in the rats during summer.
 
Under winter conditions, though, they detected various general toxicological effects housed in concrete cages. Rats caged in clay cages showed no such effects. Infrared thermographic examination indicates that skin temperature of rats in concrete cages was abnormally low, but not so in the rats in clay cages. In this study, they used 2D differential in-gel electrophoresis and tandem mass spectrometry to examine proteomic changes in the serum of the rats housed in both types of cages.
 

 
Journal: BMC Bioinformatics, Aug. 28 [Epub ahead of print]
Title: NITPICK: peak identification for mass spectrometry data
Authors: BY Renard, M Kirchner; H Steen; JA Steen: FA Hamprecht
 
Non-greed, Interative Template-based peak PICKer, or NITPICK, is proposed as a method for peak picking of features from mass spectra. NITPICK deconvolves complex overlapping isotope distributions in multi-component mass spectra and is based on fractional averagine and on a modified version of sparse, non-negative least angle regression for which a “suitable, statistically motivated early stopping criterion has been derived,” according to the abstract.
 

 
Journal: Proteomics, Aug. 19 [Epub ahead of print]
Title: High-throughput proteomic analysis of formalin-fixed paraffin-embedded tissue microarrays using MALDI imaging mass spectrometry
Authors: MR Groseclose; PP Massion; P Chaurand; RM Caprioli
 
Described is a novel method that uses on-tissue tryptic digestion followed by MALDI imaging mass spectrometry. They analyzed by mass spectrometry a tissue microarray section containing 112 needle core biopsies from lung-tumor patients. Data were correlated to a serial hematoxylin and eosin-stained section having various histological regions marked and normal ones.
 
“By correlating each mass spectrum to a defined histological region, statistical classification models were generated that can sufficiently distinguish biopsies from adenocarcinoma from squamous cell carcinoma biopsies,” according to the abstract. MALDI MS/MS sequence analysis was used to identify peptide markers directly from the tissue microarray section.
 

 
Journal: Journal of Proteome Research, Aug. 16 [Epub ahead of print]
Title: Identification of differentially expressed proteins in the cervical mucosa of HIV-1-resistant sex workers
Authors: A Burgener; J Boutilier; C Wachihi; J Kimani; M Carpenter; G Westmacott; K Cheng; TB Ball; F Plummer
 
Authors used a 2D DIGE method to examine cervical mucosa of HIV-1 resistant women for biomarkers of such resistance. They found 15 proteins to be differentially expressed greater than eight-fold. Women with resistance overexpressed several antiproteases, including those from the serpin B family, and cystatin A, a known anti-HIV-1 factor. Immunoblotting for a selection of the proteins confirmed the DIGE volume differences.
 

 
Journal: Proteomics, Aug. 14 [Epub ahead of print]
Title: Protein unlocking procedures of formalin-fixed paraffin-embedded tissues: Application to MALDI-TOF imaging MS investigations
Authors: M Ronci; E Bonanno; B Colantoni; L Pieroni; C Di Ilio; LG Spagnoli; G Federici; A Urbani
 
While archival formalin-fixed paraffin embedded tissues are a “powerful tool” for the examination of clinical course of diseases, the quality of molecular data is strongly influenced by the tissue preparation condition, according to the abstract. Authors developed an in vitro approach using “tissue surrogate” samples to explore different protein unlocking procedures that could enable the recovery of polypeptides for mass spec analysis. The results outline the possibility to obtain valuable peptide mass spectra profiles from FFPE preparations “by applying a combined two steps procedure of heat induced antigen retrieval (HIAR) in presence of EDTA and on target trypsin hydrolysis,” they said in the abstract.
 

 
Journal: Analytical Chemistry, Aug. 12 [Epub ahead of print]
Title: Assessing the dynamic range and peak capacity of nanoflow LC-FAIMS-MS on an ion trap mass spectrometer for proteomics
Authors: JD Canterbury; X Yi; MR Hoopmann; MJ MacCoss
 
Authors describe the application of a high-field asymmetric waveform ion mobility spectrometry, or FAIMS, device as an interface to an ion trap mass spectrometer. They said that adding the device to the front of the instrument allowed them to obtain a greater than eight-fold increase in peak capacity and a greater than five-fold increase in dynamic range without increasing the length of the LC-MS analysis.
 

 
Journal: Journal of Mass Spectrometry, Aug. 12 [Epub ahead of print]
Title: Oscore: a combined score to reduce false negative rates for peptide identification in tandem mass spectrometry analysis
Authors: C Shao; W Sun; F Li; R Yang; L Zhang; Y Gao
 
Despite the development of multiple algorithms for assessing matches between MS/MS spectra and peptide sequences in databases, “it is still a challenge to reduce false negative rates without compromising the high confidence of peptide identification,” the authors say in the abstract. They developed the score, Oscore, by logistic regression using SEQUEST and AMASS variables to identify fully tryptic peptides. By combining them, they improved the classification of correct and incorrect peptide identifications.
 
They report Oscore achieved a lower false negative rate and a lower false positive rate than PeptideProphet on datasets of 18 known protein mixtures and several proteome-scale samples of different complexity, database size, and separation methods.
 

 
Journal: Clinical Chemistry, Aug. 7 [Epub ahead of print]
Title: High-abundance polypeptides of the human plasma proteome comprising the top 4 logs of polypeptide abundance
Authors: GL Hortin; D Sviridov; NL Anderson
 
Authors surveyed proteomic studies to identify candidates for high-abundance polypeptide chains, and searched the literature for information on the plasma concentrations of the most abundant components in healthy adults and for the molecular mass of the mature polypeptide chains in plasma. They summarized the data on individual peptide chains for proteins containing multiple subunits or polypeptides and collected data on about 150 of the most abundant polypeptides in plasma.
 

 
Journal: Journal of the American Society for Mass Spectrometry, Aug. 7 [Epub ahead of print]
Title: How much peptide sequence information is contained in the ion trap tandem mass spectra?
Authors: J Cox, NC Hubner; M Mann
 
Authors assigned more than 400,000 identified tandem mass spectra from a single human cancer cell line project to 26,896 distinct peptide sequences. The average absolute fragment mass accuracy is 0.102 daltons. They report an average of about four complementary b- and y-ions, and half of all spectra contain uninterrupted b- and y-ion series of six or more amino-acid sequences. The sequences, they said in the abstract, are almost always unique in the human proteome “even without using any precursor or peptide sequence tag information. Thus, optimal de novo sequencing algorithms should be able to obtain substantial sequence information in at least half of all cases.”
 

 
Journal: Bioinformatics, Aug. 6 [Epub ahead of print]
Title: Peptide Finder: Mapping measure molecular masses to peptides and proteins
Authors: A Alexandridou; GT Tsangaris; K Vougas; K Nikita; G Spyrou
 
Article presents a publicly available web application “that facilitates a high resolution mapping of measured molecular masses to peptides and proteins, irrespectively of the enzyme/digestion method used,” the authors say in the abstract. The approach is complementary to other techniques for protein identification and provides insight into novel peptide discovery and protein identification when identification scores from the other approaches may be below significant threshold.

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