title: Proteogenomic analysis of Mycobacterium tuberculosis by high resolution mass spectrometry.
abstract: The genome sequencing of H37Rv strain of Mycobacterium tuberculosis was completed in 1998 followed by the whole genome sequencing of a clinical isolate, CDC1551 in 2002. Since then, the genomic sequences of a number of other strains have become available making it one of the better studied pathogenic bacterial species at the genomic level. However, annotation of its genome remains challenging because of high GC content and dissimilarity to other model prokaryotes. To this end, we carried out an in-depth proteogenomic analysis of the M. tuberculosis H37Rv strain using Fourier transform mass spectrometry with high resolution at both MS and tandem MS levels. In all, we identified 3176 proteins from Mycobacterium tuberculosis representing ~80% of its total predicted gene count. In addition to protein database search, we carried out a genome database search, which led to identification of ~250 novel peptides. Based on these novel genome search-specific peptides, we discovered 41 novel protein coding genes in the H37Rv genome. Using peptide evidence and alternative gene prediction tools, we also corrected 79 gene models. Finally, mass spectrometric data from N terminus-derived peptides confirmed 727 existing annotations for translational start sites while correcting those for 33 proteins. We report creation of a high confidence set of protein coding regions in Mycobacterium tuberculosis genome obtained by high resolution tandem mass-spectrometry at both precursor and fragment detection steps for the first time. This proteogenomic approach should be generally applicable to other organisms whose genomes have already been sequenced for obtaining a more accurate catalogue of protein-coding genes.
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore, India.
authors.forename: Dhanashree S
authors.initials: DS
authors.lastname: Kelkar
authors.forename: Dhirendra
authors.initials: D
authors.lastname: Kumar
authors.forename: Praveen
authors.initials: P
authors.lastname: Kumar
authors.forename: Lavanya
authors.initials: L
authors.lastname: Balakrishnan
authors.forename: Babylakshmi
authors.initials: B
authors.lastname: Muthusamy
authors.forename: Amit Kumar
authors.initials: AK
authors.lastname: Yadav
authors.forename: Priyanka
authors.initials: P
authors.lastname: Shrivastava
authors.forename: Arivusudar
authors.initials: A
authors.lastname: Marimuthu
authors.forename: Sridhar
authors.initials: S
authors.lastname: Anand
authors.forename: Hema
authors.initials: H
authors.lastname: Sundaram
authors.forename: Reena
authors.initials: R
authors.lastname: Kingsbury
authors.forename: H C
authors.initials: HC
authors.lastname: Harsha
authors.forename: Bipin
authors.initials: B
authors.lastname: Nair
authors.forename: T S Keshava
authors.initials: TS
authors.lastname: Prasad
authors.forename: Devendra Singh
authors.initials: DS
authors.lastname: Chauhan
authors.forename: Kiran
authors.initials: K
authors.lastname: Katoch
authors.forename: Vishwa Mohan
authors.initials: VM
authors.lastname: Katoch
authors.forename: Prahlad
authors.initials: P
authors.lastname: Kumar
authors.forename: Raghothama
authors.initials: R
authors.lastname: Chaerkady
authors.forename: Srinivasan
authors.initials: S
authors.lastname: Ramachandran
authors.forename: Debasis
authors.initials: D
authors.lastname: Dash
authors.forename: Akhilesh
authors.initials: A
authors.lastname: Pandey
mesh_terms: D000465:Algorithms; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D018834:Chaperonin 60 / Q000737:chemistry / Q000378:metabolism; D018387:Codon, Initiator; D005583:Fourier Analysis; D013058:Mass Spectrometry; D058977:Molecular Sequence Annotation; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000378:metabolism*; D016366:Open Reading Frames; D010446:Peptide Fragments / Q000737:chemistry; D021382:Protein Sorting Signals; D040901:Proteomics; D057386:Search Engine
chemical_list: D001426:Bacterial Proteins; D018834:Chaperonin 60; D018387:Codon, Initiator; D010446:Peptide Fragments; D021382:Protein Sorting Signals