Production of authentic geranylgeranylated KRAS4b using an engineered baculovirus system.

ABSTRACT

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.