Author: Adrian Lewis Guthals Publisher: ISBN: 9781321891584 Category : Languages : en Pages : 108
Book Description
High-throughput proteomics is made possible by a combination of modern mass spectrometry instruments capable of generating many millions of tandem mass (MS2 or MS/MS) spectra on a daily basis and the increasingly sophisticated associated software for their automated identification. Despite the growing accumulation of collections of identified spectra and the regular generation of MS2 data from related peptides, the mainstream approach for peptide identification is still the nearly two decades old approach of matching one MS2 spectrum at a time against a database of protein sequences. These traditional approaches fail for the identification of spectra from unknown proteins such as antibodies or proteins from organisms with un-sequenced genomes. Furthermore, attempts to identify MS/MS spectra against large databases (e.g., the human microbiome or 6-frame translation of the human genome) face a search space that is 10-100 times larger than the human proteome, where it becomes increasingly challenging to separate between true and false peptide matches. First, we describe techniques to utilize networks of spectra from related peptides to rigorously compute the joint spectral probability of multiple spectra being matched to peptides with overlapping sequences, thus improving peptide identification by 30-62% against large search spaces. We then introduce methods that dramatically improve de novo sequencing of unknown proteins using novel spectral network assembly algorithms and incorporating alternative MS/MS acquisition protocols. Finally, we describe an interesting end-goal biological problem for which the described advances in de novo sequencing can usher in a new era of therapeutic drug discovery.
Author: Michael Kinter Publisher: John Wiley & Sons ISBN: 0471231886 Category : Science Languages : en Pages : 321
Book Description
How to design, execute, and interpret experiments for protein sequencing using mass spectrometry The rapid expansion of searchable protein and DNA databases in recent years has triggered an explosive growth in the application of mass spectrometry to protein sequencing. This timely and authoritative book provides professionals and scientists in biotechnology research with complete coverage of procedures for analyzing protein sequences by mass spectrometry, including step-by-step guidelines for sample preparation, analysis, and data interpretation. Michael Kinter and Nicholas Sherman present their own high-quality, laboratory-tested protocols for the analysis of a wide variety of samples, demonstrating how to carry out specific experiments and obtain fast, reliable results with a 99% success rate. Readers will get sufficient experimental detail to apply in their own laboratories, learn about the proper selection and operation of instruments, and gain essential insight into the fundamental principles of mass spectrometry and protein sequencing. Coverage includes: * Peptide fragmentation and interpretation of product ion spectra * Basic polyacrylamide gel electrophoresis * Preparation of protein digests for sequencing experiments * Mass spectrometric analysis using capillary liquid chromatography * Techniques for protein identification by database searches * Characterization of modified peptides using tandem mass spectrometry And much more
Author: Oscar Alzate Publisher: CRC Press ISBN: 1420076264 Category : Medical Languages : en Pages : 356
Book Description
In this, the post-genomic age, our knowledge of biological systems continues to expand and progress. As the research becomes more focused, so too does the data. Genomic research progresses to proteomics and brings us to a deeper understanding of the behavior and function of protein clusters. And now proteomics gives way to neuroproteomics as we beg
Author: Rune Matthiesen Publisher: ISBN: 9781627033923 Category : Mass spectrometry Languages : en Pages : 405
Book Description
Since the publishing of the first edition, the methodologies and instrumentation involved in the field of mass spectrometry-based proteomics has improved considerably. Fully revised and expanded, Mass Spectrometry Data Analysis in Proteomics, Second Edition presents expert chapters on specific MS-based methods or data analysis strategies in proteomics. The volume covers data analysis topics relevant for quantitative proteomics, post translational modification, HX-MS, glycomics, and data exchange standards, among other topics. Written in the highly successful Methods in Molecular Biology series format, chapters include brief introductions to their respective subjects, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Updated and authoritative, Mass Spectrometry Data Analysis in Proteomics, Second Edition serves as a detailed guide for all researchers seeking to further our knowledge in the field of proteomics.
Author: Christian Earl Henry Beaudrie Publisher: ISBN: Category : Languages : en Pages : 262
Book Description
"Protein identification via tandem mass spectrometry and database searching has become a powerful tool in high-throughput proteomics. However, many tandem mass spectra are noisy, are formed by poorly fragmented peptides, and do not result in peptide assignment. Furthermore, many 'high-quality' un-assigned spectra are discarded as noise when they contain sufficient information for an assignment. A novel method was developed to assess the quality of tandem mass spectra from a LC-QTOF instrument using multivariate classification techniques. Six mass spectral features were first evaluated for their ability to discriminate between 'low-' and 'high-quality' spectra. Next, features were used in combination with three classification methods to create an optimal discriminator. Individual spectra quality scores were determined for two datasets, allowing for the removal of 'low quality' spectra, and a focused database search on 'high quality' unassigned spectra. Using these methods it was possible to reduce data size by 43.2% with a loss of only 5% of 'high-quality' spectra, while increasing peptide identifications by over 6%." --
Author: Willy Vincent Bienvenut Publisher: Springer Science & Business Media ISBN: 9781402033186 Category : Medical Languages : en Pages : 324
Book Description
At present where protein identification and characterisation using mass spectrometry is a method of choice, this book is presenting a review of basic proteomic techniques. The second part of the book is related to the novel high throughput protein identification technique called the 'molecular scanner'. Several protein identification techniques are described, especially the peptide mass fingerprint with MALDI-MS based method. E.g. ionisation process, matrix available, signal reproducibility and suppression effect, as well as date treatment for protein identification using bioinformatics tools.
Author: Conrad Bessant Publisher: Royal Society of Chemistry ISBN: 1782626735 Category : Science Languages : en Pages : 429
Book Description
The field of proteomics has developed rapidly over the past decade nurturing the need for a detailed introduction to the various informatics topics that underpin the main liquid chromatography tandem mass spectrometry (LC-MS/MS) protocols used for protein identification and quantitation. Proteins are a key component of any biological system, and monitoring proteins using LC-MS/MS proteomics is becoming commonplace in a wide range of biological research areas. However, many researchers treat proteomics software tools as a black box, drawing conclusions from the output of such tools without considering the nuances and limitations of the algorithms on which such software is based. This book seeks to address this situation by bringing together world experts to provide clear explanations of the key algorithms, workflows and analysis frameworks, so that users of proteomics data can be confident that they are using appropriate tools in suitable ways.
Author: John R. Griffiths Publisher: John Wiley & Sons ISBN: 1119045851 Category : Science Languages : en Pages : 414
Book Description
Covers all major modifications, including phosphorylation, glycosylation, acetylation, ubiquitination, sulfonation and and glycation Discussion of the chemistry behind each modification, along with key methods and references Contributions from some of the leading researchers in the field A valuable reference source for all laboratories undertaking proteomics, mass spectrometry and post-translational modification research
Author: Laurence Elliot Bernstein Publisher: ISBN: Category : Languages : en Pages : 125
Book Description
Tandem mass spectrometry (MS/MS) has long been the leading method of identifying peptides and proteins in complex biological samples and many algorithms have been created for this purpose. Many of the methods for searching MS/MS spectra against a database of known proteins must restrict the number of post-translational modifications (PTMs) that they can identify because the larger the number of PTMs being considered, the larger the search space, which in turn increases both computational complexity and the potential for false matches. In addition these algorithms cannot discover new peptides or homologues or be used with species for which a protein database does not exist. Newer algorithms have been developed that perform "open" or "blind" searches capable of finding any possible modifications, however these methods increase the search space even further, often resulting in lower performance and the generation of many putative modification masses that must be sifted through manually to determine which are real. To address the shortcomings of the existing methods, we created a new blind database search algorithm based on spectral networks. Our method uses a modification of the standard spectral tagging filtration techniques tailored for contig-consensus spectra generated from spectral networks, along with, the first of its kind, penalty-based, dynamic programming spectrum-database alignment algorithm that is able to accurately to identify both a priori specified modifications as well as novel PTMs. We then developed a workflow based on these new techniques that combines previous work in clustering, spectral alignment, spectral networks, and multi-spectral assembly. Because our new algorithm only identifies spectra that lie within the spectral networks, we created a workflow, called RaVen, that merged our method with MS-GF+ and combines the results from both methods resulting in a method with massive improvement in overall identification rates above existing methods while at the same time identifying many more rare modifications in samples. We also propose an improved way of measuring the accuracy of blind search algorithms: "peptide variants" which better meet captures the goals of blind search methods and does not rely on precise localization of modifications (which is very difficult to achieve for most algorithms).
Author: Adrian Lewis Guthals
Author: Michael Kinter
Author: 
Author: Oscar Alzate
Author: Rune Matthiesen
Author: Christian Earl Henry Beaudrie
Author: Willy Vincent Bienvenut
Author: Conrad Bessant
Author: John R. Griffiths
Author: Laurence Elliot Bernstein