RRC ID |
22783
|
著者 |
Mon H, Lee J, Fukushima M, Nagata Y, Fujii M, Xu J, Nishi O, Iiyama K, Kusakabe T.
|
タイトル |
Production and characterization of the celery mismatch endonuclease CEL II using baculovirus/silkworm expression system.
|
ジャーナル |
Appl Microbiol Biotechnol
|
Abstract |
Mutation and polymorphism detection by nucleases has become a more important tool in clinical and biological researches. There are several kinds of single-stranded nucleases for detecting mismatched DNAs. One of them, CEL II, was isolated from Apium graveolens and cleaves DNA with high specificity at sites of mismatch. High-throughput mutation scanning requires large quantity of CEL II endonuclease. Here, we demonstrate high-level expression of CEL II using silkworm-baculovirus system. The recombinant CEL II secreted in silkworm hemolymph was glycosylated and susceptible to N-glycosidase F. Additionally, larger metal ions such as Ca(2+) and Sr(2+) were able to replace Mg(2+) and enhanced mismatch cleavage activity of CEL II. These results indicate that the silkworm-baculovirus platform is a good alternative system to obtain the functional CEL II.
|
巻・号 |
97(15)
|
ページ |
6813-22
|
公開日 |
2013-8-1
|
DOI |
10.1007/s00253-012-4583-1
|
PMID |
23179626
|
MeSH |
Animals
Apium / enzymology*
Baculoviridae / genetics*
Base Sequence
Bombyx / genetics*
DNA Primers
DNA Restriction Enzymes / metabolism*
Glycosylation
|
IF |
3.53
|
引用数 |
10
|
WOS 分野
|
BIOTECHNOLOGY & APPLIED MICROBIOLOGY
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リソース情報 |
カイコ |
d17 |