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Title: Structural genomic studies of lipoproteins from Mycobacterium smegmatis for drug design
Author: Almourfi, Feras
ISNI:       0000 0004 5349 1798
Awarding Body: University of Sheffield
Current Institution: University of Sheffield
Date of Award: 2014
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Tuberculosis (TB) is considered as an old infectious disease that leads to many fatalities in Man. Mycobacterium tuberculosis was discovered as the causative agent of tuberculosis by Robert Koch in 1882. Since then, scientists started the first move in order to develop such tools to prevent the disease. According to the WHO 2012 TB report, about one-third of the population is infected with M.tuberculosis; TB causes nearly 1.8 million deaths every year. Perhaps most worrying, new strains of M.tuberculosis resistant to most or even all-standard anti-TB drugs are spreading throughout the world, making treatment more costly and often impossible. Therefore, we urgently need to discover new drugs to overcome TB. The genomic sequence of M.tuberculosis has been completed in 1998 and has helped to shed light on new pathways as drug targets. As part of a drug discovery programme, a structural genomics study of lipoproteins has been launched using the Mycobacterium smegmatis as a model organism for M.tuberculosis. A lipid-anchored protein is a class of protein that is produced in the cytoplasm as a pre-prolipoprotein and attached to the cell membrane following posttranslational modification (lipidation). Such proteins represent about 3% of bacterial genomes. Furthermore, all bacteria apparently allocate particular proteins to the cell envelope by a process called post-translational lipid modification in order to produce membrane-anchored lipoproteins that are able to work in the aqueous environment at the membrane interface. Therefore, this project aims to identify new targets suitable for drug discovery and shed light on their role in the cell. Eight targets were identified and put into a pipeline of cloning, over-expression, purification and crystallization for structure determination. Five target proteins of different putative functions were successfully purified with one (Msmeg_0515) annotated as an ABC sugar transporter protein, leading to structure determination. The structure of Msmeg_0515 (AgaE) has been determined to a high resolution of 1.22 A. Structure comparison of AgaE with other sugar binding proteins revealed that AgaE shares a similar fold with the maltose/maltodextrin binding protein (MalE) from E.coli. Previous bioinformatics studies on the sugar transporters of M.smegmatis and M.tuberculosis suggested that AgaE is an α-galactoside sugar binding protein, however, structural analysis of the binding site of AgaE protein revealed that it's more similar to malto-oligosaccharide binding proteins. Also, binding assay by Circular dichroism (CD) has revealed a significant affinity of AgaE for maltose, glycerol 3-phosphate and acarbose but not α-galactoside sugars.
Supervisor: Patrick, Baker Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available