1996 Fiscal Year Final Research Report Summary
X-ray diffraction study and reaction mechanism of bacterial alginate lyase
| Project/Area Number |
07660111
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
応用微生物学・応用生物化学
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| Research Institution | KYOTO UNIVERSITY |
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Principal Investigator |
MURATA Kousaku Research Institute for Food Science Department of Food Design and Utilization : Professor, 食糧科学研究所, 教授 (90142299)
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| Co-Investigator(Kenkyū-buntansha) |
MIKAMI Bunzo Research Institute for Food Science Department of Food Design and Utilization :, 食糧科学研究所, 助教授 (40135611)
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| Project Period (FY) |
1995 – 1996
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| Keywords | Bacterial infection / Pseudomonas aeruginosa / Alginate / X-ray diffraction / Alginate lyase |
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| Research Abstract |
Alginate is a highly viscous heteropolysaccharide composed of manuronate and gluronate. Alginate formation by Pseudomonas aeruginosa is a leading cause of bacterial infectious disease, which is also called biofilm infection. The effective therapeutic methods for the disease have not been developed. For the treatment of bacterial infectious disease, the removal of diofilm is expected as a useful and effective procedures. The present study aimed to apply bacterial enzyme : alginate lyase that depolymerizes alginate for the removal of biofilm and to analyze three-dimensional structure. Following results were obtained. (1) Preliminary X-ray diffraction study was performed and crystal constants were determined. However, the enzyme crystal did not accept heavy metal ions, although the reason for this resistance is unclear. (2) The reaction mechanism of the lyase was analyzed and reaction constants were obtained with an estimation of the active site structure. (3) The alginate lyase was chemically modified with polyyethylenglycol. The modified enzyme showed sufficient lyase activity with extremely low lebel of antigenic activity. The results, together with others, strongly suggested that the enzyme will be applied to the treatment of bacterial infectious patients. (4) To further decrease antigenic activity, the alginate lyase was fragmented and low molecular size enzyme was generated. (5) The bacterium producing alginate lyase had a pit on the cell surface. This is the first finding in the history of microbiology. (6) The fine structural analysis of the pit structure by electron microscopy and pit-deficient mutant indicated that the pit is responsible for the uptake of macromolecules with energy and information. Our paper summarized these results was awarded from Japan Society of Fermentation and Bioengineering in 1996.
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