Enhancing the thermal tolerance of bacillus subtilis ASUIA243 phytase using homology modeling and molecular dynamics simulation /
Phytases catalyze the degradation of phytic acid or phytate (IP6) to lower inositol phosphate esters and inorganic phosphate. Phytase is mainly used as a feed supplement for swine and poultry. As these animals are monogastric, they cannot metabolize phytate. Hence, phytase is added to animal feeds t...
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| Main Author: | |
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| Format: | Thesis |
| Language: | English |
| Published: |
Kuala Lumpur :
Kulliyyah of Engineering, International Islamic University Malaysia,
2012
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| Subjects: | |
| Online Access: | http://studentrepo.iium.edu.my/handle/123456789/4776 |
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| Summary: | Phytases catalyze the degradation of phytic acid or phytate (IP6) to lower inositol phosphate esters and inorganic phosphate. Phytase is mainly used as a feed supplement for swine and poultry. As these animals are monogastric, they cannot metabolize phytate. Hence, phytase is added to animal feeds to utilize the phosphorus in phytate and reduce its secretion, which causes ecological problems (eutrophication). The preparation of animal feeds involves a pelleting process that uses high temperature (60-80°C). As a result, it's crucial to have thermostable phytases to avoid loss of activity during pelleting. This study aimed at improving the thermostability of Bacillus subtilis ASUIA243 phytase using computational (in silica) methods. Homology modeling was used to develop a 3D structure of B. subtilis ASUIA243 phytase using the crystal structure of Bacillus amyloliquefaciens phytase (PDB code 1H6L) as a template, where both enzymes showed 72% sequence identity. B. subtilis ASUIA243 phytase model was validated and found to have equivalent quality with B. amyloliquefaciens phytase. Molecular dynamics (MD) simulation was used to investigate the dynamic properties of phytase to i,d. entify factors contributing to thermostability. Root Mean Square Deviation (RMSD) fluctuations of secondary structures were analyzed. Additionally, static properties such as number of hydrogen bonds and salt bridges were examined. Experimental mutation study of Aspergillus niger phytase that resulted in a mutant enzyme with 20% increase in thermostability was used as control. Three mutants were introduced to the B. subtilis ASUIA243 phytase. Analysis of RMSD fluctuations showed that, with regards to the results of wild-type and thermostable mutant A. niger phytase, mutant2 (D145N/Pl49D) was found to be the most promising among all three B. subtilis ASUIA243 mutants, as it highly resembles the dynamic behavior of the thermostable mutant A. niger phytase. In addition, it is anticipated that mutant 1 (R248T /T250E) and mutant2 (D145N/P149D) are likely to retain the activity and structural stability of B. subtilis ASUIA243 model phytase. The choice of A. niger phytase and its thermophilic mutant was not ideal for this study as the enzyme has a different fold and a small difference in biochemical properties. However, experimental mutation studies have only been conducted for E. coli and A. niger phytases, where the latter was the better choice. |
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| Item Description: | Abstracts in English and Arabic. " A dissertation submitted in fulfilment of the requirement for the degree of Master of Science (Biotechnology Engineering)."--On t.p. |
| Physical Description: | xviii, 166 leaves : illustrations ; 30cm. |
| Bibliography: | Includes bibliographical references (leaves 151-155). |