K-Ras4B lipoprotein synthesis: Biochemical characterization, functional properties, and dimer formation

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• 作者：Alexey Dementiev ; ¹ ; ^{adement@uic.edu} ; ^{seevich3326@hotmail.com}
• 关键词：Small GTPase ; K-Ras4B ; Lipoprotein ; Nanodisc ; Membrane protein ; Dimer
• 刊名：Protein Expression and Purification
• 出版年：2012
• 期刊代码：184_10465928
• 类别：bio
• 出版时间：July, 2012
• 卷：84
• 期：1
• 页码：86-93
• 文件大小：598 K

摘要

K-Ras4B, a small GTPase and a key oncogene, plays a central role in the early steps of signal transduction from activated receptor tyrosine kinases by recruiting its downstream effectors to the cell membrane. Specific posttranslational modifications of K-Ras4B, including the addition of C-terminal farnesyl and methyl groups, mediate its proper membrane localization and signaling activity. The mechanism and molecular determinants underlying this selective membrane localization and molecular interactions with its many regulators and downstream effectors are largely unknown. Preparative amounts of the posttranslationally processed K-Ras4B protein are necessary to carry out structural, functional, and cell biological studies of this important oncogene. In this work we describe a simple and efficient method for synthesis of milligram quantities of functionally active, fully processed K-Ras4B. Using this preparation, we observe K-Ras4B dimerization in vitro; this has not been observed previously and could be important for its activity, membrane anchoring, and translocation between different cellular membranes.