DESIGN AND IDENTIFICATION OF NETWORK METABOLIC REACTION IN MYCOBACTERIUM

TUBERCULOSIS BY SYSTEM BIOLOGY APPRAOCH

PLAN OF STUDY:

Mycobacterium tuberculosis continues to be a major pathogen in the third world, killing almost 2
million people a year by the most recent estimates. Even in industrialized countries, the
emergence of multi-drug resistant (MDR) strains of tuberculosis hails the need to develop
additional medications for treatment. Many of the drugs used for treatment of tuberculosis target
metabolic enzymes. Genome-scale models can be used for analysis, discovery, and as hypothesis
generating tools, which will hopefully assist the rational drug development process. These
models need to be able to assimilate data from large datasets and analyze them.

Tuberculosis continues to be a devastating pathogen throughout the world,

particularly in developing nations. In 2001, the World Health Organization (WHO) estimated 8.5
million new cases of tuberculosis (based on 3.8 million new reported cases) and an estimated 1.8
million deaths from tuberculosis in 2000 . Within the United States, the number of reported cases
of tuberculosis has been decreasing with the exception of a period when the trend reversed in
1986 and peaked in 1992 .This reversal has been attributed principally to HIV/AIDs,
immigration from countries with high prevalence of tuberculosis, poverty, homelessness, and
multi-drug resistant (MDR) tuberculosis .MDR tuberculosis is generally defined as strains that
are resistant to treatment with isoniazid and rifampin , two of the key first line antituberculosis
drugs . MDR strains of tuberculosis emerged in the early 1990s and have now been found all
over the world .

Many of the unique properties of tuberculosis are attributable to its metabolism,

particularly the complex fatty acids characteristics of the organism. These mycolic acids,
phenolic glycolipids, and mycoceric acids confer many of the properties such as its acid-fastness
and are believed to contribute to the resilience of the organism. Mycobacterium tuberculosis can
survive in a wide range of environments (many different tissues) and fairly extreme pHs. One of
the most confounding factors with these bacteria is their ability to survive for long periods of
time in a dormant stage. The slow doubling time of tuberculosis has further limited the amount
of experimental data that can be generated. Many of the first and second line drugs used to treat
tuberculosis have metabolic targets, so developing systems level models of metabolism are
anticipated to be of great use in the future.

The purpose of this research is to obtain a comprehensive list of all pathways

known so far in literature. We are planned to identify the Mtb proteins for which no reaction
information exists in KEGG.and many important for solving some biological problems . The
challenge is to mine reaction information from literature and other published reports in Cell
Designer using SBML. This will be followed by validating reactions reported in KEGG and
creating the network of metabolic reactions in Mtb.
Research steps:

Phase 1
You are all assigned the Glycolytic pathway as a test case

We know from standard biochemistry text books/ literature the sequence of reactions in this
pathway . The task at hand is to identify the proteins in M.tuberculosis (by their GI/Rv numbers)
and annotate them in detail. You will all independently try to obtain data for the first four
proteins in the Glycolysis pathway, namely, Hexokinase, Phosphoglucose isomerase,
Phosphofructokinase and Aldolase, and fill in the details as per the indicated fields in above
form.

a. Identify the correct protein from Mycobacterium tuberculosis H37Rv genome (Available as a
resource)

b. The fields can be filled by referring to any of the publicly available databases such as
TubercuList (http://genolist.pasteur.fr/Tuberculist or http://tuberculist.epfl.ch), NCBI
(http://www.ncbi.nlm.nih.gov; RefSeq ID : NC_000962), Sanger
(http://www.sanger.ac.uk/Projects/M_tuberculosis/), KEGG (http://www.genome.jp/kegg/),
BioCyc (http://biocyc.org/MTBRV/)

Form Fields and their Definitions

 Rv ID (Identifiers of the ORFs in Mycobacterium tuberculosis H37Rv, e.g., Rv2763c)

 Gene name (Corresponding gene name of the Rv number, e.g., dfrA, folA)
 Pathway name (Pathway to which the Rv number belongs to, e.g., Folate biosynthesis)
 Annotation (Name of the gene product, e.g., Dihydrofolate reductase)
 Reaction substrate(s) (Substrates involved in the reaction, e.g., 5,6,7,8‐Tetrahydrofolate +
NADP(+))
 Reaction Product(s) (Products formed in the reaction, e.g., 7,8‐Dihydrofolate + NADPH)
 EC number (Enzyme Commission number; available only if the protein is an enzyme,
e.g., 1.5.1.3)
 Function (Function of the protein/gene product, e.g., Essential step for de novo glycine
and purine synthesis, DNA precursor synthesis, and for the conversion of dUMP to
dTMP)

Phase II
Your Task from 21st December to 28th December 2009

From the above task you will get all the information to create reactions in Cell Designer. Use
information generated from the steps and follow Cell Designer tutorial (attached below) to create
pathway in SBML format. You are requested to upload the SBML files to our group.

IMPORTANT: Before uploading the SBML file to our group please validate them.

Phase III
Please go through the list of the pathways below and search literature to see if there are any
pathways (IN ANY SPECIES) that are missing from the list. Any such pathway that you notice,
you may add to the list, along with your name in the field provided. Also, please cite the
references for each entry, in the field provided for that. These pathways can belong to any
species, can be metabolic/signaling or regulatory. This activity has no deadline as of now.
List of Pathway From KEGG Suggest New Pathways (Not available in KEGG)
Reference:

1. Design and construction of the platform for comparative genomics.Liu N, Guo YB, Sun
XH, Ma L, Deng QK Nan Fang Yi Ke Da Xue Xue Bao. 2010 Feb;30(2):219-23.
2. Analysis of the genetic variation in Mycobacterium tuberculosis strains by multiple
genome alignments Andrés Cubillos-Ruiz,1 Juan Morales,2 and María Mercedes
Zambrano BMC Res Notes. 2008; 1: 110.
Genomic analysis reveals variation between Mycobacterium tuberculosis H37Rv and the
attenuated M. tuberculosis H37Ra strain.Brosch R, Philipp WJ, Stavropoulos E, Colston
MJ, Cole ST, Gordon SV. Infect Immun 2000 Jan;68(1):427.