Cells and cell lysates: a direct approach for engineering antibodies against membrane proteins using yeast surface display.
Cells and cell lysates: a direct approach for engineering antibodies against membrane proteins using yeast surface display.
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细胞和细胞裂解物：利用酵母表面展示工程化抗膜蛋白抗体的直接方法。
DOI：
10.1016/j.ymeth.2012.03.010
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发表时间：
2013
期刊：
影响因子：
--
通讯作者：
Shusta,EricV
中科院分区：
文献类型：
--
作者：
Tillotson,BenjaminJ;Cho,YongKu;Shusta,EricV
膜蛋白通常是抗体工程的理想靶点。然而，大多数抗体工程平台隐含地依赖于目标抗原的水溶性，这通常是MPs的问题。重组、可溶形式的MPs已成功地用作抗体工程的抗原来源，但可溶性MPs片段的异种表达和纯化仍然是一个具有挑战性和耗时的过程。在这里，我们提出了一种更直接的方法来帮助MPs抗体的工程设计。通过将酵母表面展示技术直接与全细胞或洗涤剂溶解的全细胞裂解物相结合，可以针对MP抗原的接近天然构象筛选抗体文库。我们还描述了该平台如何适用于抗体表征和抗原鉴定。这一系列相容的方法为MPs抗体工程提供了基础，预计这些方法将随着膜蛋白生物化学和增溶技术的发展而成熟。
Membrane proteins (MPs) are often desirable targets for antibody engineering. However, the majority of antibody engineering platforms depend implicitly on aqueous solubility of the target antigen which is often problematic for MPs. Recombinant, soluble forms of MPs have been successfully employed as antigen sources for antibody engineering, but heterologous expression and purification of soluble MP fragments remains a challenging and time-consuming process. Here we present a more direct approach to aid in the engineering of antibodies to MPs. By combining yeast surface display technology directly with whole cells or detergent-solubilized whole-cell lysates, antibody libraries can be screened against MP antigens in their near-native conformations. We also describe how the platform can be adapted for antibody characterization and antigen identification. This collection of compatible methods serves as a basis for antibody engineering against MPs and it is predicted that these methods will mature in parallel with developments in membrane protein biochemistry and solubilization technology.