PROJECT REPORTON

Homology Modeling of Mycobacterium tuberculosis H37Rv gyra Protein :Design and Evaluation of novel gyrA inhibitors

Submitted in partial fulfillment of the requirements forthe award of degree of

BACHELOR OF TECHNOLOGY

IN

BIOINFORMATICS

By

Anurag YadavMRT07UGBBI009

Under the supervision of

Prof. Dwijendra K. Gupta Ms.Sarika SahuCo-ordinator, Bioinformatics Assistant ProfessorDepartment of Biochemistry School of BiotechnologyUniversity of Allahabad Center of Bioinformatics

Session: 2007 2011

SHOBHIT UNIVERSITYModipuram Meerut-250110

School of Biotechnology &Centre for Bioinformatics

Shobhit University, Meerut

CertificateThis is to certify that the work embodied in the project report entitled HomologyModeling of Mycobacterium tuberculosis H37Rv gyrA Protein : Design andEvaluation of novel gyrA inhibitors has been carried out by Mr. Anurag Yadavunder our supervision for the partial fulfillment of the requirements for the degree ofBachelor of Technology in Biotechnology.

(Prof. D.V. Rai)Director

(Ms. Sarika Sahu)Supervisor

DECLARATION

I Anurag Yadav, confirm that this work submitted for assessment is my originalwork and is expressed in my own words. Any uses made within it of the works ofother authors in any form (e.g. ideas, equations, figures, text, tables,programmes) are properly acknowledged at the point of their use. A full list ofthe references employed has been included.I also declare that this project report has not been submitted any where in anyform for the partial fulfillment of the award of degree.I understand that my project report may be made electronically to the public.

(Anurag Yadav)

Abstract

The present study focuses on determining and validating the structure of a protein target andconducting a preliminary in silico study on Trovofloxacin and Moxifloxacin compounds for thedesign of a putative drug.

gyrA encodes Dna gyrase subunit A, important for cell DNA replication andmultiplication of bacteria. The crystal structure of gyrA is not available in any structural database.In this study we modeled a 3D structure of gyrA by X-ray crystal structure of N-terminal domain ofDna gyrase subunit A of Mycobcterium tuberculosis H37Rv (PDB ID: 3IFZ, Chain A), used as thetemplate.

The receptor was docked to the gyrA inhibitor drugs and final energy value of each confomerobtained as follows Gatifloxacin (-11.2837), Ciprofloxacin (-11.2814), Norfloxacin (-9.7557),Moxifloxacin (-11.3729), Trovafloxacin (-11.4397), Gemifloxacin (-11.0109) and Tamifloxacin (-8.4628) using the docking suite of MOE. Depending on the energy values the best two drugs, they areTrovafloxacin and Moxifloxacin are chosen. We tried to improve the binding efficiency and stericcompatibility of these two drugs namely Trovafloxacin and Moxifloxacin.

Several modifications were made to the probable functional groups which were interacting with thereceptor molecule. Analogs of these drug molecule were prepared using ACD ChemSketch anddocked using MOE docking . Trovafloxacin Analog 6 and Moxifloxacin analog 6 were detected withsignificant energy values.

ACKNOWLEDGEMENT

First and foremost, I would like to thank my supervisor, Mrs. Sarika Sahu, for his guidance,support and for sharing his knowledge with me throughout my B.tech and Mr. Surya PratapSingh and Mrs. Nutan Chauhan, for proof-reading my thesis and for the great enthusiasm andmotivation that every meeting with her brought..

I am thankful to Prof. Anoop Swarup Vice Chancellor, Shobhit University,Meerut, for providing all the necessary requirements and for his moral support for thisdissertation work as well during the whole course of B. Tech. I am also thankful to Prof. R.P.Agarwal, Pro-vice chancellor Shobhit University, Meerut for his honest dedication towards oureducation and career and for being with us in various levels of academic pursuits.

In this project, I was greatly assisted, inspired by Prof. Dwijendra Guptawithout them, the completion of this project was almost impossible. With great reverence, Iacknowledge them for providing me an environment to involve as an independent researcher.Their constant encouragement and affection gave me courage to tackle the failure days, which isinevitable in a researchers life.

My primary thanks goes to Surya Pratap Singh and Ansul Tiwari for hisdeep commitment & guidance. I am also grateful to Dr. Rekha Dixit, Mr. Maneesh Gupta andMr.Vikas Gupta for their motivation throughout my project work. It is my pleasure to thank allthe people who helped to turn this project a success.

Contents

1 : Introduction1

1.1 Motivation1.2 Problem Statement1.3 Introduction to Trovafloxacin1.4 Introduction to Moxifloxacin1.5 Target of Trovafloxacin and Moxifloxacin1.6 Dna gyrase

2 : Literature review.4

2.1 Intoduction to Mycobacterium genome2.1.1 Disease caused by Mycobaacterium tuberculosis2.1.2 Sign and Symptom

2.2 Computer aided drug design2.2.1 Introduction2.2.2 Drug design cycle

2.3 Homology modeling2.3.1 General Procedures

2.4 Docking

3: Materials and Tools.12

3.1 : ACD/ChemSkecth3.2 : MOE(Molecular Operating Enviroment)3.3 : Drug Bank3.4 : PDB3.5 : Pubmed3.6 : Modeller 9v7

4: Methodology.14

4.1 : Screening for best homologoues templates

4.2 : In silico Comprative Modelling of gyrA protein

4.3 : Model refinement, validation and evaluation

4.4 : Submission of Modeled structure to PMDB

4.5 : Selection of potential drug candidate against gyrA

4.6 : Ligand

4.7 : Docking in MOE

4.7.1 : Methodology Overview

5: Result22

5.1 : Docking result with Trovafloxacin Derivative

5.2 : Docking result with Moxifloxacin Derivative

6: Discussion.35

7: Conclusion36

8: Future work..37

9: References..38

List of figures

Figure 1: Structure of Trovafloxacin

Figure 2: Structure of Moxifloxacin

Figure 3: M.tuberculosis colonies

Figure 4: Symptoms of M.tuberculosis in Human lung

Figure 5: 3-D structure of gyrA protein modeled by Modeller 9v7

Figure 6: Ramachandran plot of target protein

List of tables

Table 1: Structure of Trovafloxacin derivatives

Table 2: Structure of Moxifloxacin derivatives

Table 3: Docking result with Trovafloxacin derivatives

Table 4: Docking result with Moxifloxacin derivatives

Abbreviations

gyrA : Dna gyrase subunit A

RMSD : Root Mean Square Deviation

E_conf : Energy of Conformer

S : Final Energy

PDB : Protein Data Bank

PMDB : Protein Model Database

MW : Molecular Weight

H-Bond : Hydrogen Bond

H-acc : Hydrogen Bond Acceptor

H-don : Hydrogen Bond donor

T.B : Tuberculosis

MDR : Multi Drug Resistance

MTB : Mycobacterium tuberculosis