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Issue 15, 2011
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trans,trans-2,4-Hexadiene incorporation on enzymes for site-specific immobilization and fluorescent labeling

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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 °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.

Graphical abstract: trans,trans-2,4-Hexadiene incorporation on enzymes for site-specific immobilization and fluorescent labeling

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Publication details

The article was received on 14 Mar 2011, accepted on 26 Apr 2011 and first published on 27 Apr 2011


Article type: Paper
DOI: 10.1039/C1OB05401E
Org. Biomol. Chem., 2011,9, 5535-5540

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    trans,trans-2,4-Hexadiene incorporation on enzymes for site-specific immobilization and fluorescent labeling

    M. Filice, O. Romero, J. M. Guisan and J. M. Palomo, Org. Biomol. Chem., 2011, 9, 5535
    DOI: 10.1039/C1OB05401E

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