Glyoxyl agarose:: A fully inert and hydrophilic support for immobilization and high stabilization of proteins
Glyoxyl agarose:: A fully inert and hydrophilic support for immobilization and high stabilization of proteins
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DOI：
10.1016/j.enzmictec.2005.10.014
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发表时间：
2006-06-26
影响因子：
3.4
通讯作者：
Guisan, Jose M.
中科院分区：
文献类型：
--
作者：
Mateo, Cesar;Palomo, Jose M.;Guisan, Jose M.
乙醛酰琼脂糖由含有大量非常稳定的醛基的相当厚的琼脂糖纤维构成，所述醛基通过非常短的间隔臂连接到支持物。在碱性条件下，这些活化的载体通过至少涉及具有较高氨基密度的蛋白质表面区域的两点反应来支持蛋白质。这些键是弱席夫碱，并且键的可逆性已被用于将该基质转化为色谱基质。通过在合适的条件下长期孵育蛋白质-载体缀合物，可以进一步促进固定化蛋白质和活化载体之间更强烈的多点附着，而催化活性的损失最小。琼脂糖-蛋白质缀合物制备的终点是非常温和的硼氢化物还原。还原后，酶通过非常稳定的仲氨基键（与以前的伯氨基键具有非常相似的物理性质）与载体保持连接，载体上剩余的醛基转化为完全惰性的羟基。许多酶和蛋白质的非常活性和高度稳定的衍生物已经使用这些载体制备。这些蛋白质固定化方案的主要特点在这里进行了讨论。(c)2005年爱思唯尔公司All rights reserved.
Glyoxyl agarose is constituted by quite thick agarose fibres containing a large number of very stable aldehyde groups attached to the support by very short spacer arms. Under alkaline conditions, these activated supports immobilize proteins, via, at least, a two-point reaction involving the region/s of the protein surface with the higher densities of amino groups. These bonds are weak Schiff's bases and the reversibility of the bonds has been used to convert this matrix into a chromatographic one. A more intense multipoint attachment between the immobilized protein and the activated support can be further promoted, with minimal loss of catalytic activity, by a long-term incubation of the protein-support conjugate under suitable conditions. The end-point of the preparation of agarose-protein conjugates is a very mild borohydride reduction. After that reduction, the enzyme remains attached to the support by means of very stable secondary amino bonds (with very similar physical properties to those of the former primary amino ones) and the remaining aldehyde groups on the support are converted into fully inert hydroxyl groups. Very active and highly stabilized derivatives of many enzymes and proteins have been prepared using these supports. The main features of these protein immobilization protocols are discussed here. (c) 2005 Elsevier Inc. All rights reserved.