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One step purification–immobilization of fuculose-1-phosphate aldolase, a class II DHAP dependent aldolase, by using metal-chelate supports

Paper ID Volume ID Publish Year Pages File Format Full-Text
18745 42741 2006 6 PDF Available
Title
One step purification–immobilization of fuculose-1-phosphate aldolase, a class II DHAP dependent aldolase, by using metal-chelate supports
Abstract

His-tagged recombinant fuculose-1-phosphate aldolase (FucA) from E. coli has been purified by immobilized metal-chelate affinity chromatography (IMAC) at gram scale. During this operation, there was a metal exchange between FucA and the affinity matrix, being the purification yields dependent on the metal nature, which was bound to affinity matrix.One step purification–immobilization of FucA has been carried out on metal-chelate support. The preparation of a FucA immobilized derivative, available to be used as catalyst in aldol addition reactions, has been accomplished in a single step starting from E. coli cell extracts. The best results were obtained with high density support containing Co2+. The immobilization yield was 100% and the immobilized derivative showed 63% of FucA activity initially offered to the support.The best derivative of immobilized FucA is 21-fold more stable than the soluble FucA in DMF/buffer (1:4) at 25 °C and it catalyzes aldol addition between S-Cbz-Alaninal and DHAP.

Keywords
Fuculose-1-phosphate aldolase (FucA); DHAP dependent aldolase; Enzyme immobilization; Immobilized metal-chelate affinity chromatography (IMAC); Metal–enzyme; One step purification–immobilization of enzymes
First Page Preview
One step purification–immobilization of fuculose-1-phosphate aldolase, a class II DHAP dependent aldolase, by using metal-chelate supports
Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 39, Issue 1, 1 June 2006, Pages 22–27
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering