High yield expression of recombinant pro-resilin:: Lactose-induced fermentation in E coli and facile purification
High yield expression of recombinant pro-resilin:: Lactose-induced fermentation in E coli and facile purification
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DOI：
10.1016/j.pep.2006.11.003
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发表时间：
2007-03-01
影响因子：
1.6
通讯作者：
Lyons, Russell
中科院分区：
文献类型：
--
作者：
Kim, Misook;Elvin, Christopher;Lyons, Russell
弹性蛋白是一种具有优异材料性能的弹性蛋白：高回弹性和非常高的疲劳寿命。我们对生产类树脂蛋白感兴趣，这种蛋白可以通过光化学交联形成橡胶状生物材料，用于各种工业和医疗应用。已经开发出一种在小型发酵设备中生产可溶性重组蛋白的方法，使用甘油分批进行初始生长，并在碳源耗尽时通过IPTG进行初级诱导，然后在乳糖诱导培养中进行新生长。重组recr -resilin[1]在宿主菌株Escherichia coli BL21(DE3)pLysS中以高达300mg/l的水平过表达，相对于在LB培养基中通过传统IPTG诱导获得的体积生产力增加了20倍以上。在新鲜培养基中，在乳糖诱导之前进行初级诱导，在相同条件下，与未进行初级诱导相比，体积生产力和细胞特异性产量提高了2.5至3倍。该方法适用于可溶性类树脂蛋白的大规模生产，操作成本低。此外，一种简单的“盐沉淀和热净化”方法允许对大量可溶性重组树脂样蛋白进行快速高效的下游处理。这些方法将有助于研究类树脂蛋白的结构和功能特性，以及开发高弹性生物材料。(c) 2006爱思唯尔公司版权所有。
Resilin is an elastic protein with outstanding material properties: high resilience and a very high fatigue lifetime. We are interested in the production of resilin-like proteins which can be photo-chemically cross-linked to form rubbery biomaterials to be used in a variety of industrial and medicinal applications. A method has been developed for producing soluble recombinant proteins in small scale fermentation equipment using glycerol batch for initial growth and primary induction by IPTG at carbon source depletion, followed by new growth in lactose-induced culture. Recombinant recl-resilin [1] has been over-expressed in the host strain Escherichia coli BL21(DE3)pLysS at a level of up to 300mg/l, a greater than 20-fold increase in volumetric productivity, relative to that obtained from conventional IPTG induction in LB medium. The primary induction step before lactose induction in fresh medium resulted in a 2.5- to 3-fold increase of both volumetric productivity and cell specific yield compared to that without primary induction under the same conditions. This method is amenable and suitable for large scale production of soluble resilin-like proteins at a low operating cost. In addition, a simple 'salt precipitation and heat purification' method allowed rapid and efficient downstream processing of a large quantity of soluble recombinant resilin-like proteins. These methods will enable investigation of the structural and functional properties of resilin-like proteins, and the development of highly resilient biomaterials. (c) 2006 Elsevier Inc. All rights reserved.