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Immobilization and stabilization of horseradish peroxidase isoforms
Author(s) -
Husain Shahid,
Jafri Farahdiba,
Saleemuddin M.
Publication year - 1996
Publication title -
iubmb life
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.132
H-Index - 113
eISSN - 1521-6551
pISSN - 1521-6543
DOI - 10.1080/15216549600201472
Subject(s) - horseradish peroxidase , chemistry , periodate , sepharose , sodium periodate , glycosyl , covalent bond , amino acid , ethylenediamine , cationic polymerization , biochemistry , chromatography , organic chemistry , enzyme
Abstract Purified anionic and cationic isoforms of horseradish peroxidase (HRP) immobilized by coupling the amino acid side‐chain amino groups and/or carbohydrate moieties to Sepharose have been studied for their resistance to denaturation. The isoforms were treated with periodate followed by ethylenediamine to generate additional amino groups in the glycosyl residues. The immobilized preparations were: Preparation I (Sp‐aHRP, Sp‐cHRP), in which HRP was covalently immobilized via side‐chain amino groups exclusively; Preparation II (Sp‐NHaHRP, Sp‐NHcHRP), in which periodate and ethylenediamine‐treated HRP was covalently immobilized via side‐chain amino groups and amino groups incorporated into glycosyl residues. HRP isoforms in preparation II lacked about 33‐55 % carbohydrate. Both strategies of immobilization induced significant stabilization against denaturation. Inclusion of 2 mM calcium enhanced isoenzyme stability significantly.