CHARACTERIZATION OF RECOMBINANT HUMAN ALDOLASE-B AND PURIFICATION BY METAL CHELATE CHROMATOGRAPHY
CHARACTERIZATION OF RECOMBINANT HUMAN ALDOLASE-B AND PURIFICATION BY METAL CHELATE CHROMATOGRAPHY
复制标题
DOI：
10.1006/bbrc.1995.1128
复制
发表时间：
1995-01-26
影响因子：
3.1
通讯作者：
TOLAN, DR
中科院分区：
文献类型：
--
作者：
DOYLE, SA;TOLAN, DR
重组人醛缩酶B与从人肝脏中纯化的天然酶在大小、电荷、结构、果糖-1,6-二（磷酸）和果糖-1-磷酸的K-m常数以及两种底物的活性比上均相同。因此，重组醛缩酶B是天然酶的有效模型，可用于研究导致遗传性果糖不耐受的突变或在活性位点设计的其他突变。在重组酶的氨基端添加6个组氨酸残基不会改变其结构或功能特征，并且可以通过固定化金属亲和层析进行纯化。该纯化方案不需要稳定的或活性的酶，将有利于突变醛缩酶B酶的研究，否则将难以纯化。(C)学术出版社
Recombinant human aldolase B and the native enzyme purified from human liver were found to be identical in size, charge, structure, K-m constants for fructose-1,6-bis(phosphate) and fructose-1-phosphate, and the activity ratio of the two substrates. Thus recombinant aldolase B is a valid model for the native enzyme and can be used to study mutations that cause hereditary fructose intolerance or others designed in the active site. Addition of six histidine residues to the amino-terminus of the recombinant enzyme did not alter its structural or functional characteristics and allowed for purification by immobilized metal affinity chromatography. This purification protocol does not require a stable or active enzyme and will facilitate the study of mutant aldolase B enzymes that would otherwise be difficult to purify. (C) Academic Press, Inc.