Methods for Studying Interactions of Detergents and Lipids with α-Helical and β-Barrel Integral Membrane Proteins

Q1 Biochemistry, Genetics and Molecular Biology
S. Saif Hasan, Danas Baniulis, Eiki Yamashita, Mariya V. Zhalnina, Stanislav D. Zakharov, Jason T. Stofleth, William A. Cramer
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引用次数: 2

Abstract

Methods for studying interactions of protein with lipids and detergents are described for representatives of two major classes of membrane proteins: (1) the α-helical hetero-oligomeric integral cytochrome b6f complex of oxygenic photosynthesis from cyanobacteria, and (2) the outer membrane β-barrel proteins BtuB and OmpF from Gram-negative Escherichia coli bacteria. Details are presented on the use of detergents for purification and crystallization of the b6f complex as well as a method for lipid exchange. The positions of detergent and lipid molecules, which define eight potential lipid-binding sites in the b6f complex, are described. Differences in detergent strategies for isolation and crystallization of β-barrel proteins relative to those for oligomeric helical membrane proteins are discussed, and purification and assessment of protein quality by circular dichroism (CD) is presented. Curr. Protoc. Protein Sci. 74:29.7.1-29.7.30. © 2013 by John Wiley & Sons, Inc.

Abstract Image

Abstract Image

洗涤剂和脂类与α-螺旋和β-桶状整体膜蛋白相互作用的研究方法
本文介绍了两大类膜蛋白的研究方法:(1)蓝藻氧合作用的α-螺旋异聚寡聚整体细胞色素b6f复合体,(2)革兰氏阴性大肠杆菌的外膜β-桶状蛋白BtuB和OmpF。详细介绍了使用洗涤剂纯化和b6f络合物的结晶,以及脂质交换的方法。描述了洗涤剂和脂质分子的位置,它们定义了b6f复合物中8个潜在的脂质结合位点。讨论了β-桶状蛋白与低聚螺旋膜蛋白分离和结晶的洗涤策略的差异,并介绍了用圆二色性(CD)纯化和评价蛋白质质量的方法。咕咕叫。Protoc。蛋白质科学,74:29.7.1-29.7.30。©2013 by John Wiley &儿子,Inc。
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来源期刊
Current Protocols in Protein Science
Current Protocols in Protein Science Biochemistry, Genetics and Molecular Biology-Biochemistry
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期刊介绍: With the mapping of the human genome, more and more researchers are exploring protein structures and functions in living organisms. Current Protocols in Protein Science provides protein scientists, biochemists, molecular biologists, geneticists, and others with the first comprehensive suite of protocols for this growing field.
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