by Filice, Marco, Romero, Oscar, Guisan, Jose M and Palomo, Jose M
Abstract:
Lipase B from Candida antarctica (CAL-B) has been site-directedly modified by the introduction of a trans,trans-hexadiene moiety onto lipase molecules, identified by MALDI-TOF. This modification on CAL-B permitted its immobilization on Q-Sepharose supports in excellent yields (>95%) when native lipase was not immobilized at pH 7 and 25 textdegreeC. After the entire modification procedure, the catalytic activity of the protein on the solid support was surprisingly increased 2-fold. A tailor-made maleimide-fluorophore derivative was specifically covalently linked to the protein in high yield via a selective Diels-Alder reaction in aqueous media. Furthermore, the NBD-labeled-CAL-B was also immobilized on the ionic support, retaining around 80% of the specific activity. The preparation of this labeled-CAL-B was also possible by a Diels-Alder reaction on solid phase in excellent yields.
Reference:
trans,trans-2,4-Hexadiene incorporation on enzymes for site-specific immobilization and fluorescent labeling. (Filice, Marco, Romero, Oscar, Guisan, Jose M and Palomo, Jose M), In Organic & biomolecular chemistry, The Royal Society of Chemistry, volume 9, 2011.
Bibtex Entry:
@article{Filice:2011iw,
author = {Filice, Marco and Romero, Oscar and Guisan, Jose M and Palomo, Jose M},
title = {{trans,trans-2,4-Hexadiene incorporation on enzymes for site-specific immobilization and fluorescent labeling.}},
journal = {Organic {&} biomolecular chemistry},
year = {2011},
volume = {9},
number = {15},
pages = {5535--5540},
month = aug,
publisher = {The Royal Society of Chemistry},
affiliation = {Departamento de Biocat{'a}lisis, Instituto de Cat{'a}lisis (CSIC), c/marie curie 2, Cantoblanco, Campus UAM, 28049 Madrid, Spain.},
doi = {10.1039/c1ob05401e},
pmid = {21695340},
language = {English},
rating = {0},
date-added = {2016-12-12T13:04:25GMT},
date-modified = {2020-07-09T13:27:50GMT},
abstract = {Lipase B from Candida antarctica (CAL-B) has been site-directedly modified by the introduction of a trans,trans-hexadiene moiety onto lipase molecules, identified by MALDI-TOF. This modification on CAL-B permitted its immobilization on Q-Sepharose supports in excellent yields (>95%) when native lipase was not immobilized at pH 7 and 25 {textdegree}C. After the entire modification procedure, the catalytic activity of the protein on the solid support was surprisingly increased 2-fold. A tailor-made maleimide-fluorophore derivative was specifically covalently linked to the protein in high yield via a selective Diels-Alder reaction in aqueous media. Furthermore, the NBD-labeled-CAL-B was also immobilized on the ionic support, retaining around 80% of the specific activity. The preparation of this labeled-CAL-B was also possible by a Diels-Alder reaction on solid phase in excellent yields.},
url = {http://xlink.rsc.org/?DOI=c1ob05401e},
uri = {url{papers3://publication/doi/10.1039/c1ob05401e}}
}