title: Moving from unsequenced to sequenced genome: reanalysis of the proteome of Leishmania donovani.
abstract: The kinetoplastid protozoan parasite, Leishmania donovani, is the causative agent of kala azar or visceral leishmaniasis. Kala azar is a severe form of leishmaniasis that is fatal in the majority of untreated cases. Studies on proteomic analysis of L. donovani thus far have been carried out using homology-based identification based on related Leishmania species (L. infantum, L. major and L. braziliensis) whose genomes have been sequenced. Recently, the genome of L. donovani was fully sequenced and the data became publicly available. We took advantage of the availability of its genomic sequence to carry out a more accurate proteogenomic analysis of L. donovani proteome using our previously generated dataset. This resulted in identification of 17,504 unique peptides upon database-dependent search against the annotated proteins in L. donovani. These peptides were assigned to 3999 unique proteins in L. donovani. 2296 proteins were identified in both the life stages of L. donovani, while 613 and 1090 proteins were identified only from amastigote and promastigote stages, respectively. The proteomic data was also searched against six-frame translated L. donovani genome, which led to 255 genome search-specific peptides (GSSPs) resulting in identification of 20 novel genes and correction of 40 existing gene models in L. donovani.
  
  
  CONCLUSIONS
  Leishmania donovani genome sequencing was recently completed, which permitted us to use a proteogenomic approach to map its proteome and to carry out annotation of it genome. This resulted in mapping of 50% (3999 proteins) of L. donovani proteome. Our study identified 20 novel genes previously not predicted from the L. donovani genome in addition to correcting annotations of 40 existing gene models. The identified proteins may help in better understanding of stage-specific protein expression profiles in L. donovani and to identify novel stage-specific drug targets in L. donovani which could be used in the treatment of leishmaniasis. This article is part of a Special Issue entitled: Trends in Microbial Proteomics.
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India; Bioinformatics Centre, School of Life Sciences, Pondicherry University, Puducherry 605014, India.
authors.forename: Raja Sekhar
authors.initials: RS
authors.lastname: Nirujogi
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India; Rajiv Gandhi University of Health Sciences, Bangalore 560041, India.
authors.forename: Harsh
authors.initials: H
authors.lastname: Pawar
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India; Department of Biotechnology, Amrita Vishwa Vidyapeetham, Kollam 690525, India.
authors.forename: Santosh
authors.initials: S
authors.lastname: Renuse
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India.
authors.forename: Praveen
authors.initials: P
authors.lastname: Kumar
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India; Manipal University, Madhav Nagar, Manipal 576104, India.
authors.forename: Sandip
authors.initials: S
authors.lastname: Chavan
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India; Manipal University, Madhav Nagar, Manipal 576104, India.
authors.forename: Gajanan
authors.initials: G
authors.lastname: Sathe
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India; Manipal University, Madhav Nagar, Manipal 576104, India.
authors.forename: Jyoti
authors.initials: J
authors.lastname: Sharma
authors.affiliation: National Centre for Cell Sciences, Pune 411007, India.
authors.forename: Sweta
authors.initials: S
authors.lastname: Khobragade
authors.affiliation: National Centre for Cell Sciences, Pune 411007, India.
authors.forename: Janhavee
authors.initials: J
authors.lastname: Pande
authors.affiliation: National Centre for Cell Sciences, Pune 411007, India.
authors.forename: Bhakti
authors.initials: B
authors.lastname: Modak
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India; Bioinformatics Centre, School of Life Sciences, Pondicherry University, Puducherry 605014, India; Manipal University, Madhav Nagar, Manipal 576104, India.
authors.forename: T S Keshava
authors.initials: TS
authors.lastname: Prasad
authors.affiliation: Institute of Bioinformatics, International Technology Park, Bangalore 560066, India.
authors.forename: H C
authors.initials: HC
authors.lastname: Harsha
authors.affiliation: National Centre for Cell Sciences, Pune 411007, India. Electronic address: patole@nccs.res.in.
authors.forename: Milind S
authors.initials: MS
authors.lastname: Patole
authors.affiliation: McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore 21205, MD, USA; Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore 21205, MD, USA; Department of Oncology, Johns Hopkins University School of Medicine, Baltimore 21205, MD, USA; Department of Pathology, Johns Hopkins University School of Medicine, Baltimore 21205, MD, USA. Electronic address: pandey@jhmi.edu.
authors.forename: Akhilesh
authors.initials: A
authors.lastname: Pandey
mesh_terms: D030562:Databases, Protein; D017125:Genes, Protozoan / Q000502:physiology*; D006801:Humans; D007893:Leishmania donovani / Q000235:genetics* / Q000378:metabolism; D007898:Leishmaniasis, Visceral / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000235:genetics* / Q000378:metabolism; D020543:Proteome / Q000235:genetics* / Q000378:metabolism; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D010455:Peptides; D020543:Proteome; D015800:Protozoan Proteins
keywords: Digenic parasite; FDR; False discovery rate; GSSPs; Genome annotation; Genome search specific peptides; HCD; High energy collision induced dissociation; High resolution mass spectrometry; Intracellular pathogen; PSM; Peptide spectrum matches