title: HCV Envelope protein 2 sequence comparison of Pakistani isolate and In-silico prediction of conserved epitopes for vaccine development.
abstract: BACKGROUND
  HCV is causing hundreds of cases yearly in Pakistan and has become a threat for Pakistani population. HCV E2 protein is a transmembrane protein involved in viral attachment and thus can serve as an important target for vaccine development but because of its variability, vaccine development against it has become a challenge. Therefore, this study was designed to isolate the HCV E2 gene from Pakistani HCV infected patients of 3a genotype, to perform In-silico analysis of HCV E2 isolated in Pakistan and to analyze HCV E2 protein sequence in comparison with other E2 proteins belonging to 3a and 1a genotypes to find potential conserved B-cells and T-cell epitopes that can be important in designing novel inhibitory compounds and peptide vaccine against genotype 3a and 1a.
  
  
  METHODS
  Patients were selected on the basis of elevated serum ALT and AST levels at least for six months, histological examination, and detection of serum HCV RNA anti-HCV antibodies (3rd generation ELISA). RNA isolation, cDNA synthesis, amplification, cloning and sequencing was performed from 4 patient's serum samples in order to get the HCV E2 sequence. HCV E2 protein of Pakistani origin was analyzed using various bioinformatics tools including sequence and structure tools.
  
  
  RESULTS
  HCV E1 protein modeling was performed with I-TASSER online server and quality of the model was assessed with ramchandran plot and Z-score. A total of 3 B-cell and 3 T-cell epitopes were found to be highly conserved among HCV 3a and 1a genotype.
  
  
  CONCLUSIONS
  The present study revealed potential conserved B-cell and T-cell epitopes of the HCV E2 protein along with 3D protein modeling. These conserved B-cell and T-cell epitopes can be helpful in developing effective vaccines against HCV and thus limiting threats of HCV infection in Pakistan.
authors: Human Molecular Biology Group, Department of Bioinformatics and Biotechnology, Government College University (GCU), Faisalabad, Pakistan.
authors: Sobia
authors: S
authors: Idrees
authors: Usman A
authors: UA
authors: Ashfaq
authors: Saba
authors: S
authors: Khaliq
mesh_terms: D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D019295:Computational Biology / Q000379:methods; D017124:Conserved Sequence; D017931:DNA Primers; D018076:DNA, Complementary / Q000378:metabolism; D004220:Disulfides; D000939:Epitopes / Q000737:chemistry*; D018985:Epitopes, B-Lymphocyte / Q000276:immunology; D018984:Epitopes, T-Lymphocyte / Q000276:immunology; D005838:Genotype; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010154:Pakistan; D017433:Protein Structure, Secondary; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000235:genetics
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D004220:Disulfides; D000939:Epitopes; D018985:Epitopes, B-Lymphocyte; D018984:Epitopes, T-Lymphocyte; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus
references: Rev Med Virol. 2000 Mar-Apr;10(2):101-17
references: 10713597
references: Virol J. 2011 Apr 11;8:161
references: 21477382
references: Virol J. 2013 Apr 10;10:113
references: 23575359
references: J Mol Recognit. 2008 Jul-Aug;21(4):243-55
references: 18496882
references: Proc Natl Acad Sci U S A. 1996 Mar 5;93(5):1759-63
references: 8700831
references: EMBO J. 2002 Oct 1;21(19):5017-25
references: 12356718
references: BMC Bioinformatics. 2008 Jan 23;9:40
references: 18215316
references: J Gen Virol. 2004 Jun;85(Pt 6):1497-1507
references: 15166434
references: J Med Virol. 1999 Mar;57(3):223-9
references: 10022791
references: Nucleic Acids Res. 2007 Jul;35(Web Server issue):W407-10
references: 17517781
references: Science. 1998 Oct 30;282(5390):938-41
references: 9794763
references: J Virol. 2005 Jul;79(13):8400-9
references: 15956584
references: Curr Opin Microbiol. 2000 Oct;3(5):445-50
references: 11050440
references: Nucleic Acids Res. 2010 Jan;38(Database issue):D854-62
references: 19906713
references: Immunity. 2010 Oct 29;33(4):530-41
references: 21029963
references: BMC Bioinformatics. 2007 Jan 05;8:4
references: 17207271
references: Proc Natl Acad Sci U S A. 2003 Apr 15;100(8):4498-503
references: 12676990
references: J Transl Med. 2011 Nov 10;9:194
references: 22074322
references: J Biol Chem. 2005 Mar 25;280(12):11329-39
references: 15611113
references: J Virol. 2001 Dec;75(24):12088-97
references: 11711599
references: J Hepatol. 1991;13 Suppl 4:S6-14
references: 1668332
references: Theor Biol Med Model. 2013 Apr 10;10:24
references: 23575038
references: J Microbiol Immunol Infect. 2008 Feb;41(1):4-8
references: 18327420
references: Nucleic Acids Res. 2005 Jul 1;33(Web Server issue):W230-2
references: 15980459
references: Genet Vaccines Ther. 2011 Apr 06;9:7
references: 21466709
references: J Mol Biol. 2001 Jan 19;305(3):567-80
references: 11152613
references: J Exp Med. 2003 Mar 3;197(5):633-42
references: 12615904
references: J Virol. 1997 Jan;71(1):697-704
references: 8985401
references: Methods Mol Biol. 1999;112:531-52
references: 10027275
references: BMC Infect Dis. 2008 May 23;8:69
references: 18498666
references: BMC Bioinformatics. 2006 Mar 13;7:131
references: 16533401
references: Cell Mol Life Sci. 2008 Jan;65(1):100-12
references: 17914604
references: J Transl Med. 2011 Jul 18;9:112
references: 21762538