Summary: Top-down mass spectrometry enables the observation of whole complex proteoforms in biological samples and provides crucial information complementary to bottom-up spectrometry. Because complexity top-down spectra proteoforms, it is a challenging problem efficiently interpret tandem high-throughput proteome-level proteomics studies. We present TopPIC, tool that identifies characterizes with unknown primary structure alterations, such as amino acid mutations post-translational...

Capillary zone electrophoresis (CZE)-tandem mass spectrometry (MS/MS) has been recognized as a useful tool for top-down proteomics. However, its performance deep proteomics is still dramatically lower than widely used reversed-phase liquid chromatography (RPLC)-MS/MS. We present an orthogonal multidimensional separation platform that couples size exclusion (SEC) and RPLC based protein prefractionation to CZE-MS/MS of Escherichia coli. The generated high peak capacity (∼4000) intact proteins,...

Capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry (CZE-ESI-MS/MS) has been recognized as an invaluable platform for top-down proteomics. However, the scale of proteomics using CZE-MS/MS is still limited due to low loading capacity and narrow separation window CZE. In this work, first time we systematically evaluated dynamic pH junction method focusing intact proteins during CZE-MS. The optimized junction-based approached a 1 μL capacity, 90 min window, high peak...

Native proteomics aims to characterize complex proteomes under native conditions and ultimately produces a full picture of endogenous protein complexes in cells. It requires novel analytical platforms for high-resolution liquid-phase separation prior mass spectrometry (MS) MS/MS. In this work, size-exclusion chromatography (SEC)-capillary zone electrophoresis (CZE)-MS/MS was developed discovery mode, resulting the identification 144 proteins, 672 proteoforms, 23 from Escherichia coli...

Labeling approaches using isobaric chemical tags (e.g., tagging for relative and absolute quantification, iTRAQ tandem mass tag, TMT) have been widely applied the quantification of peptides proteins in bottom-up MS. However, until recently, successful applications these to top-down proteomics limited because tend precipitate "crash" out solution during TMT labeling complex samples making such difficult. In this study, we report a MS platform confidently identifying quantifying low molecular...

There are two approaches for de novo protein sequencing: Edman degradation and mass spectrometry (MS). Existing MS-based methods characterize a novel by assembling tandem spectra of overlapping peptides generated from multiple proteolytic digestions the protein. Because each spectrum covers only short peptide target protein, key to high coverage sequencing is find spectral pairs in order assemble long ones. However, regions may be too confidently identified. High-resolution spectrometers...

Although proteomics has rapidly developed in the past decade, researchers are still early stage of exploring world complex proteoforms, which protein products with various primary structure alterations resulting from gene mutations, alternative splicing, post-translational modifications, and other biological processes. Proteoform identification is essential to mapping proteoforms their functions as well discovering novel new functions. Top-down mass spectrometry method choice for identifying...

Various proteoforms may be generated from a single gene due to primary structure alterations (PSAs) such as genetic variations, alternative splicing, and post-translational modifications (PTMs). Top-down mass spectrometry is capable of analyzing intact proteins identifying patterns multiple PSAs, making it the method choice for studying complex proteoforms. In top-down proteomics, proteoform identification often performed by searching tandem spectra against protein sequence database that...

We study a bio-detection application as case to demonstrate that Kmeans -- based unsupervised feature learning can be simple yet effective alternative deep techniques for small data sets with limited intra-as well inter-class diversity. investigate the effect on classifier performance of augmentation extraction multiple patch sizes and at different image scales. Our set includes 1833 images from four classes bacteria, each bacterial culture captured three wavelengths overall collected during...

We study a bio-detection application as case to demonstrate that Kmeans -- based unsupervised feature learning can be simple yet effective alternative deep techniques for small data sets with limited intra-as well inter-class diversity. investigate the effect on classifier performance of augmentation extraction multiple patch sizes and at different image scales. Our set includes 1833 images from four classes bacteria, each bacterial culture captured three wavelengths overall collected during...

Capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry (CZE-ESI-MS/MS) has been recognized as a useful tool for top-down proteomics that aims to characterize proteoforms in complex proteomes. However, the application of CZE-MS/MS large-scale impeded by low sample-loading capacity and narrow separation window CZE. Here, protocol is described using with microliter-scale volume 90-min proteomics. The platform based on linear polyacrylamide (LPA)-coated capillary...

Top-down mass spectrometry plays an important role in intact protein identification and characterization. spectra are more complex than bottom-up because they often contain many isotopomer envelopes from highly charged ions, which may overlap with one another. As a result, spectral deconvolution, converts top-down spectrum into monoisotopic list, is key step interpretation. In this paper, we propose new scoring function, L-score, for evaluating envelopes. By combining L-score MS-Deconv,...

Abstract Complex proteoforms contain various primary structural alterations resulting from variations in genes, RNA, and proteins. Top‐down mass spectrometry is commonly used for analyzing complex because it provides whole sequence information of the proteoforms. Proteoform identification by top‐down spectral database search a challenging computational problem types and/or locations some target are general unknown. Although alignment graph algorithms have been proposed identifying with...

Top-down mass spectrometry has unique advantages in identifying proteoforms with multiple post-translational modifications and/or unknown alterations. Most software tools this area search top-down spectra against a protein sequence database for proteoform identification. When the species studied experiment lacks its proteome database, homologous can be used The accuracy of sequences affects sensitivity identification and shift localization. We tested TopPIC, commonly tool spectral...

Database search is the main approach for identifying proteoforms using top-down tandem mass spectra. However, it extremely slow to align a query spectrum against all protein sequences in large database when target proteoform that produced contains post-translational modifications and/or mutations. As result, efficient and sensitive sequence filtering algorithms are essential speeding up search. In this paper, we propose novel algorithm, which generates graphs from subspectra of searches them...

Top-down mass spectrometry is capable of identifying whole proteoform sequences with multiple post-translational modifications because it generates tandem spectra directly from intact proteoforms. Many software tools, such as ProSightPC, MSPathFinder, and TopMG, have been proposed for proteoforms modifications. In these various methods are employed to estimate the statistical significance identifications. However, most existing designed identifications without modifications, challenge...

Abstract Although proteomics has made rapid progress in the past decade, researchers are still early stage of exploring world complex proteoforms, which protein products with various primary structure alterations resulting from gene mutations, alternative splicing, post-translational modifications, and other biological processes. Proteoform identification is essential to mapping proteoforms their functions as well discovering novel new functions. Top-down mass spectrometry method choice for...

The method of physical fingerprint spectrum for Reduning injection (RI) was proposed in this paper to improve its quality standards based on the strong correlation between physicochemical properties drugs, their safety, effectiveness and stability. RI studied by thought chemistry. visually showed radar map, consisted eight indexes (pH, conductivity, turbidity, refractive index, osmolarity, surface tension, relative density, kinematic viscosity). Then 12 batch samples were verified. It found...

The rapid development of next generation sequencing technology is changing the way biological research in many aspects, which has become most popular platform for genomic structural variation detection.In this paper, we focus on single-sample data analysis, and propose a hierarchical structure to model dispersion minor allele frequency genome scale.The empirical Bayes method employed estimate hyper-parameters, identified as error or heterozygous according posterior probabilities.We suggest...