Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman
Tuberculosis (TB) infection is well-known for millennia and has been well apprehended since the end of 19th century. Robert Koch discovered its causative agent, Mycobacterium tuberculosis in 1882. TB vaccine has been in used for about a century, and antibiotics have been in place and utilized for...
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| Format: | Thesis |
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2017
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| Online Access: | http://studentsrepo.um.edu.my/8362/ http://studentsrepo.um.edu.my/8362/2/All.pdf http://studentsrepo.um.edu.my/8362/6/maikudi.pdf |
| _version_ | 1848773639124549632 |
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| author | Mohammed Maikudi , Usman |
| author_facet | Mohammed Maikudi , Usman |
| author_sort | Mohammed Maikudi , Usman |
| building | UM Research Repository |
| collection | Online Access |
| description | Tuberculosis (TB) infection is well-known for millennia and has been well apprehended
since the end of 19th century. Robert Koch discovered its causative agent, Mycobacterium
tuberculosis in 1882. TB vaccine has been in used for about a century, and antibiotics
have been in place and utilized for more than six decades. They all work, yet 1.5 million
deaths were recorded in 2014 due to TB which made it the leading infectious killer disease
globally. Bacillus Calmette-Guérin (BCG), a live attenuated vaccine and is currently the
most widely used. It has variability in effectiveness ranging from 0-80%, hence an urgent
need for a better vaccine candidate is of paramount important. The recombinant clone
(pET30a / Myt272-3 clone) constructed in Molecular Bacteriology and Toxicology
Laboratory of University of Malaya was screened for its stability, and was found stable.
The recombinant clone was transformed into BL21 (DE3) pLysS strain of Escherichia
coli. The Myt272-3 protein was successfully expressed in pET30a / Myt272-3 clone. The
molecular weight of the protein was found to be approximately 10.58 kDa as determined
by SDS-PAGE and conformed to the MW computed by Expert Protein Analysis System
(EXPASY MW bioinformatics tool). Protein BLAST (Basic Local Alignment Search
tool) bioinformatics analysis indicated 81% homology with Phenolpthiocerol synthesis
polyketide synthase I PpSA of Mycobacterium tuberculosis, MALDI-TOF analysis
further validated the homology of the protein. The concentration of protein was
determined by detergent-compatible method of protein assay. The protein was purified
by both Nickel based (Nickel-nitrilotriacetic acid, Ni-NTA) and Cobalt based
(Dynabeads®) affinity chromatographic techniques. Recently, computational biology
approaches are found very useful for organizing and understanding huge data leading to
the new field called immunoinformatics. Bioinformatics software were used to analyze the protein sequence for predictions of allergenicity, antigenicity, major
histocompatibility complexes, I and II binding and B-cell epitope binding. Moreover,
toxicity of epitopes was predicted via toxicity predictive tool. These predictive findings
serve as a practical guide towards Mycobacterium tuberculosis peptide vaccine design
and development. |
| first_indexed | 2025-11-14T13:45:36Z |
| format | Thesis |
| id | um-8362 |
| institution | University Malaya |
| institution_category | Local University |
| last_indexed | 2025-11-14T13:45:36Z |
| publishDate | 2017 |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | um-83622020-05-31T22:24:49Z Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman Mohammed Maikudi , Usman Q Science (General) R Medicine (General) Tuberculosis (TB) infection is well-known for millennia and has been well apprehended since the end of 19th century. Robert Koch discovered its causative agent, Mycobacterium tuberculosis in 1882. TB vaccine has been in used for about a century, and antibiotics have been in place and utilized for more than six decades. They all work, yet 1.5 million deaths were recorded in 2014 due to TB which made it the leading infectious killer disease globally. Bacillus Calmette-Guérin (BCG), a live attenuated vaccine and is currently the most widely used. It has variability in effectiveness ranging from 0-80%, hence an urgent need for a better vaccine candidate is of paramount important. The recombinant clone (pET30a / Myt272-3 clone) constructed in Molecular Bacteriology and Toxicology Laboratory of University of Malaya was screened for its stability, and was found stable. The recombinant clone was transformed into BL21 (DE3) pLysS strain of Escherichia coli. The Myt272-3 protein was successfully expressed in pET30a / Myt272-3 clone. The molecular weight of the protein was found to be approximately 10.58 kDa as determined by SDS-PAGE and conformed to the MW computed by Expert Protein Analysis System (EXPASY MW bioinformatics tool). Protein BLAST (Basic Local Alignment Search tool) bioinformatics analysis indicated 81% homology with Phenolpthiocerol synthesis polyketide synthase I PpSA of Mycobacterium tuberculosis, MALDI-TOF analysis further validated the homology of the protein. The concentration of protein was determined by detergent-compatible method of protein assay. The protein was purified by both Nickel based (Nickel-nitrilotriacetic acid, Ni-NTA) and Cobalt based (Dynabeads®) affinity chromatographic techniques. Recently, computational biology approaches are found very useful for organizing and understanding huge data leading to the new field called immunoinformatics. Bioinformatics software were used to analyze the protein sequence for predictions of allergenicity, antigenicity, major histocompatibility complexes, I and II binding and B-cell epitope binding. Moreover, toxicity of epitopes was predicted via toxicity predictive tool. These predictive findings serve as a practical guide towards Mycobacterium tuberculosis peptide vaccine design and development. 2017-09 Thesis NonPeerReviewed application/pdf http://studentsrepo.um.edu.my/8362/2/All.pdf application/pdf http://studentsrepo.um.edu.my/8362/6/maikudi.pdf Mohammed Maikudi , Usman (2017) Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman. PhD thesis, University of Malaya. http://studentsrepo.um.edu.my/8362/ |
| spellingShingle | Q Science (General) R Medicine (General) Mohammed Maikudi , Usman Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman |
| title | Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman |
| title_full | Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman |
| title_fullStr | Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman |
| title_full_unstemmed | Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman |
| title_short | Protein analysis and characterization of recombinant clone Myt272-3 towards tuberculosis vaccine / Mohammed Maikudi Usman |
| title_sort | protein analysis and characterization of recombinant clone myt272-3 towards tuberculosis vaccine / mohammed maikudi usman |
| topic | Q Science (General) R Medicine (General) |
| url | http://studentsrepo.um.edu.my/8362/ http://studentsrepo.um.edu.my/8362/2/All.pdf http://studentsrepo.um.edu.my/8362/6/maikudi.pdf |