| RRC ID |
22783
|
| Author |
Mon H, Lee J, Fukushima M, Nagata Y, Fujii M, Xu J, Nishi O, Iiyama K, Kusakabe T.
|
| Title |
Production and characterization of the celery mismatch endonuclease CEL II using baculovirus/silkworm expression system.
|
| Journal |
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.
|
| Volume |
97(15)
|
| Pages |
6813-22
|
| Published |
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
|
| Times Cited |
10
|
|
WOS Category
|
BIOTECHNOLOGY & APPLIED MICROBIOLOGY
|
| Resource |
| Silkworms |
d17 |
