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Construction of metagenome libraries and characterization of a novel amylolytic enzyme : 메타게놈 라이브러리의 구축과 발견된 신규 아밀로라이틱 효소의 특성 규명
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- Authors
- Advisor
- 유상렬
- Issue Date
- 2003
- Publisher
- 서울대학교 대학원
- Description
- Thesis (master`s)--서울대학교 대학원 :농생명공학부,2003.
- Abstract
- The classical approach for isolating enzymes from environmental
samples that includes enrichment of microorganisms in pure culture can
analyze only subset of the total microbiota in nature since it has been
estimated that less than 1% of microorganisms in nature can be cultivated
by standard techniques. Metagenome was isolated directly from two
different soil samples and metagenomic libraries were constructed using
pUC19 vector to obtain useful microbial enzymes from uncultured soil
microorganisms. The libraries were screened for amylase, cellulase,
protease, DNase and lipase. Initial screening revealed that one clone from
approximately 12,000 recombinant Escherichia coli strains had amylase
activity. Sequencing of the clone revealed that the amylolytic enzyme was
expressed from a novel gene. The putative amylase gene was
overexpressed and purified for characterization. The amylase had 42˚C
and pH 9.5 as the optimum temperature and pH conditions. Ca2+ was
found to be the essential element for enzyme stability. The amylase
hydrolyzed soluble starch, β-cyclodextrin to produce high level of
maltose and pullulan was also hydrolyzed to produce panose. The enzyme showed high transglycosylation activity to make 1,4-linkages exclusively.
These results suggest that the amylase is a novel enzyme conferring
maltose producing α-amylase activity.
- Language
- English
- URI
- http://dcollection.snu.ac.kr:80/jsp/common/DcLoOrgPer.jsp?sItemId=000000058889
https://hdl.handle.net/10371/67611
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