Procter Lauren, Grose Carissa, Esposito Dominic
Protein Expression Laboratory, NCI RAS Initiative, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc. PO Box B, Frederick, MD, 21702, USA.
Protein Expression Laboratory, NCI RAS Initiative, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc. PO Box B, Frederick, MD, 21702, USA.
Protein Expr Purif. 2018 Nov;151:99-105. doi: 10.1016/j.pep.2018.06.012. Epub 2018 Jun 22.
Protein prenylation is a vital eukaryotic post-translational modification which permits interaction of proteins with cellular membranes. Prenylated proteins are involved in a number of human diseases, and play a major role in cancers driven by the oncogene KRAS, which is normally farnesylated. In cases where the farnesylation machinery is inhibited, however, KRAS eludes inactivation by using an alternative prenylation pathway in which the protein is geranylgeranylated. In order to study this alternative prenylation, large quantities of accurately processed protein are required. We have developed a system to permit high-yield production of geranylgeranylated KRAS which utilizes an engineered baculovirus system. The development of this system helped to elucidate a potential metabolic bottleneck in insect cell production that should enable better production of any geranylgeranylated proteins using this system.
蛋白质异戊二烯化是一种重要的真核生物翻译后修饰，它使蛋白质能够与细胞膜相互作用。异戊二烯化的蛋白质与多种人类疾病有关，并且在由致癌基因KRAS驱动的癌症中起主要作用，KRAS通常会被法尼基化。然而，在法尼基化机制受到抑制的情况下，KRAS会通过使用另一种异戊二烯化途径来逃避失活，在这种途径中蛋白质会被香叶基香叶基化。为了研究这种替代的异戊二烯化，需要大量精确加工的蛋白质。我们开发了一种系统，该系统利用工程化杆状病毒系统实现香叶基香叶基化KRAS的高产。该系统的开发有助于阐明昆虫细胞生产中潜在的代谢瓶颈，这应该能够使使用该系统更好地生产任何香叶基香叶基化的蛋白质。
