通过阳离子弹性蛋白样多肽改进重组蛋白的非色谱纯化。
Improved non-chromatographic purification of a recombinant protein by cationic elastin-like polypeptides.
作者信息
Lim Dong Woo, Trabbic-Carlson Kimberly, Mackay J Andrew, Chilkoti Ashutosh
机构信息
Department of Biomedical Engineering, Box 90281, Duke University, Durham, North Carolina 27708-0281, USA.
出版信息
Biomacromolecules. 2007 May;8(5):1417-24. doi: 10.1021/bm060849t. Epub 2007 Apr 4.
Abstract
This paper reports an improvement in the purification of thioredoxin (Trx) expressed from E. coli by inverse transition cycling (ITC) using cationic elastin-like polypeptides (ELPs). Two ELP libraries having 2% and 5% lysine residues and molecular weights ranging from 4 to 61.1 kDa showed greater salt sensitivity in their inverse transition behavior than purely aliphatic ELPs. Expression yield of Trx-ELP fusions was an unpredictable function of guest residue composition, but reducing the molecular weight of the ELP tag generally increased Trx yield. A cationic 4.3 kDa ELP is the shortest ELP used to purify any protein by ITC to date. A 15.9 kDa ELP with a guest residue composition of K:V:F of 1:7:1 was found to be the optimal cationic tag to purify Trx, as it provided 50% greater Trx yield and only required one-fifth the added NaCl for purification of Trx as compared to previously used aliphatic ELP tags.
摘要
本文报道了使用阳离子弹性蛋白样多肽（ELP）通过反向转变循环（ITC）对大肠杆菌表达的硫氧还蛋白（Trx）进行纯化的改进。两个赖氨酸残基含量分别为2%和5%、分子量在4至61.1 kDa之间的ELP文库，其反向转变行为比纯脂肪族ELP表现出更高的盐敏感性。Trx-ELP融合蛋白的表达产量是客体残基组成的不可预测的函数，但降低ELP标签的分子量通常会提高Trx产量。阳离子4.3 kDa ELP是迄今为止通过ITC纯化任何蛋白质所使用的最短的ELP。发现客体残基组成为K:V:F = 1:7:1的15.9 kDa ELP是纯化Trx的最佳阳离子标签，因为与先前使用的脂肪族ELP标签相比，它使Trx产量提高了50%，并且纯化Trx所需添加的NaCl仅为其五分之一。
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