Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins

The NS1A protein is a non-structural protein from influenza A virus H1N1 strain. The protein is a multifunctional protein which is capable of blocking the defense mechanism of host immune by inhibiting the secretion of host cell IFN a/ß. Even existing vaccines cannot protect host cells against this...

Full description

Saved in:

Bibliographic Details
Main Author:	Chelliah, Kumutha
Format:	Thesis
Language:	English
Published:	2012
Subjects:	Q Science (General) QH301 Biology
Online Access:	http://eprints.utm.my/id/eprint/36995/5/KumuthaChelliahMFBB2012.pdf
Tags:	Add Tag No Tags, Be the first to tag this record!

id	my-utm-ep.36995
record_format	uketd_dc
spelling	my-utm-ep.369952017-09-30T01:53:47Z Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins 2012-07 Chelliah, Kumutha Q Science (General) QH301 Biology The NS1A protein is a non-structural protein from influenza A virus H1N1 strain. The protein is a multifunctional protein which is capable of blocking the defense mechanism of host immune by inhibiting the secretion of host cell IFN a/ß. Even existing vaccines cannot protect host cells against this viral infection due to constant mutations of NS1A protein. In this study, the NS1A gene which was formerly cloned in pET 32c(+) vector was successfully mutated using error-prone PCR with increased concentration of MgCl2 to 10 mM and subsequently cloned into yT&A vector and transformed into E. coli DH5a. There were four proteins that contain non-conservative mutations from sequencing which were NS1 F103LN209D, NS1 S7P, NS1 T76I and NS1 E159G mutant proteins. These proteins together with the wild-type protein were modeled using EasyModeller 2.1 and were energy minimized using GROMACS. The qualities of the structures were validated using ERRAT, PROCHECK, Verify3D and ProSA web. All the structures were of good quality and the high RMSD value shows that the mutant proteins have low structural homology to the wild-type protein. This proves that the structures were affected by point mutations. None of the mutations fell into ‘hot spot’ mutations. These proteins were subsequently docked to RNA aptamers via HEX server to analyze the binding regions and binding affinity of aptamers to proteins. The results obtained shows that the protein mutations affect the binding properties of aptamers to the mutant proteins because aptamers were docked at various regions with different binding affinities. The aptamers with the highest binding affinity towards wild-type NS1A protein and mutant proteins were selected which were aptamers 21, 174 and 176. These results were expected to be useful for potential drug design to curb future H1N1 viral infections. 2012-07 Thesis http://eprints.utm.my/id/eprint/36995/ http://eprints.utm.my/id/eprint/36995/5/KumuthaChelliahMFBB2012.pdf application/pdf en public masters Universiti Teknologi Malaysia, Faculty of Biosciences and Bioengineering Faculty of Biosciences and Bioengineering
institution	Universiti Teknologi Malaysia
collection	UTM Institutional Repository
language	English
topic	Q Science (General) QH301 Biology
spellingShingle	Q Science (General) QH301 Biology Chelliah, Kumutha Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins
description	The NS1A protein is a non-structural protein from influenza A virus H1N1 strain. The protein is a multifunctional protein which is capable of blocking the defense mechanism of host immune by inhibiting the secretion of host cell IFN a/ß. Even existing vaccines cannot protect host cells against this viral infection due to constant mutations of NS1A protein. In this study, the NS1A gene which was formerly cloned in pET 32c(+) vector was successfully mutated using error-prone PCR with increased concentration of MgCl2 to 10 mM and subsequently cloned into yT&A vector and transformed into E. coli DH5a. There were four proteins that contain non-conservative mutations from sequencing which were NS1 F103LN209D, NS1 S7P, NS1 T76I and NS1 E159G mutant proteins. These proteins together with the wild-type protein were modeled using EasyModeller 2.1 and were energy minimized using GROMACS. The qualities of the structures were validated using ERRAT, PROCHECK, Verify3D and ProSA web. All the structures were of good quality and the high RMSD value shows that the mutant proteins have low structural homology to the wild-type protein. This proves that the structures were affected by point mutations. None of the mutations fell into ‘hot spot’ mutations. These proteins were subsequently docked to RNA aptamers via HEX server to analyze the binding regions and binding affinity of aptamers to proteins. The results obtained shows that the protein mutations affect the binding properties of aptamers to the mutant proteins because aptamers were docked at various regions with different binding affinities. The aptamers with the highest binding affinity towards wild-type NS1A protein and mutant proteins were selected which were aptamers 21, 174 and 176. These results were expected to be useful for potential drug design to curb future H1N1 viral infections.
format	Thesis
qualification_level	Master's degree
author	Chelliah, Kumutha
author_facet	Chelliah, Kumutha
author_sort	Chelliah, Kumutha
title	Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins
title_short	Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins
title_full	Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins
title_fullStr	Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins
title_full_unstemmed	Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins
title_sort	random mutagenesis of ns1 protein of influenza a h1n1 and docking of rna aptamers to wild type and mutant ns1 proteins
granting_institution	Universiti Teknologi Malaysia, Faculty of Biosciences and Bioengineering
granting_department	Faculty of Biosciences and Bioengineering
publishDate	2012
url	http://eprints.utm.my/id/eprint/36995/5/KumuthaChelliahMFBB2012.pdf
_version_	1747816489548775424

Random mutagenesis of NS1 protein of influenza a H1N1 and docking of RNA aptamers to wild type and mutant NS1 proteins

Similar Items