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Efficient isolation of carbonyl‐reducing enzymes using affinity approach with anticancer drug oracin as a specific ligand
Author(s) -
Škarydová Lucie,
Andrýs Rudolf,
Holubová Lucie,
Štambergová Hana,
Kňavová Jana,
Wsól Vladimír,
Bílková Zuzana
Publication year - 2013
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.201201141
Subject(s) - chemistry , enzyme , biotransformation , ligand (biochemistry) , combinatorial chemistry , covalent bond , isoquinoline , xenobiotic , drug metabolism , ligand efficiency , alkylation , stereochemistry , biochemistry , organic chemistry , receptor , catalysis
Carbonyl‐reducing enzymes are important in both metabolism of endogenous substances and biotransformation of xenobiotics. Because sufficient amounts of native enzymes must be obtained to study their roles in metabolism, an efficient purification strategy is very important. Oracin (6‐[2‐(2‐hydroxyethyl)aminoethyl]‐5,11‐dioxo‐5,6‐dihydro‐11 H ‐indeno[1,2‐ c ] isoquinoline) is a prospective anticancer drug and one of the xenobiotic substrates for carbonyl‐reducing enzymes. A new purification strategy based on molecular recognition of carbonyl‐reducing enzymes with oracin as a ligand is reported here. The type of covalent bond, ligand molecules orientation, and their distance from the backbone of the solid matrix for good stearic accessibility were taken into account during the designing of the carrier. The carriers based on magnetically active microparticles were tested by recombinant enzymes AKR 1 C 3 and CBR 1. The S i MAG ‐ COOH magnetic microparticles with N ‐alkylated oracin and BAPA as spacer arm provide required parameters: proper selectivity and specificity enabling to isolate the target enzyme in sufficient quantity, purity, and activity.