Post-translational modification of proteins by 15-carbon and 20-carbon isoprenoids in three mammalian cell lines

Abstract

A number of cellular proteins, including p21ras, lamin B, and the G-protein gamma subunits, undergo post-translational modification by 15-carbon farnesyl or 20-carbon geranylgeranyl isoprenoid moieties derived from pyrophosphate intermediates of the cholesterol biosynthetic pathway. In this study, isoprenylated proteins in three mammalian cell lines (Hela cells, Rat-6 fibroblasts and COS cells) were radiolabeled with an isoprenoid precursor, [3H]mevalonate, and resolved by SDS gel electrophoresis. Groups of proteins with different molecular masses were eluted from the gels and the chain-lengths of the radiolabeled isoprenyl groups, released from the proteins by Raney-nickel-catalyzed desulfurization, were established by gel permeation chromatography. 15-Carbon and 20-carbon isoprenyl groups were found in separate classes of proteins within each cell line. With the exception of p21ras, which incorporated a 15-carbon group when expressed in COS cells, the proteins in the region of the 21-28 kDa ras-related GTP binding proteins contained mostly 20-carbon isoprenyl chains. In contrast, proteins belonging to the 66-72 kDa nuclear lamin family, as well as unidentified proteins with molecular masses of 41-46 kDa and 53-55 kDa, contained predominantly 15-carbon isoprenyl chains. The chain-lengths of the isoprenoids associated with particular classes of proteins did not vary from one cell line to another, suggesting that the nature of the isoprenoid modification (farnesyl versus geranylgeranyl) is determined by intrinsic structural features of the proteins, rather than the cell type in which the proteins are expressed.

Molecular cloning and characterization of the genes encoding the proteins of Zika virus

“…Lamin A is a nuclear protein and connected to the nuclear membrane by an isoprenyl group that is associated with membrane through protein-protein interaction (Reese and Maltese, 1991). Lamin A is required for early biological activities of cell mitosis by depolymerization of lamins (Reese and Maltese, 1991). During mitosis, duplicated centrosomes separate and form mitotic spindles with microtubules.…”

Identification and Characterization of a Calmodulin-binding Domain in Ral-A, a Ras-related GTP-binding Protein Purified from Human Erythrocyte Membrane

“…Ral proteins undergo sequential posttranslational modifications that occur at a CAAL motif in the C-terminal region of the molecules (38). Ral-A translation products in reticulocyte lysates have been found to be modified by 20-carbon isoprenyl groups (39). Very recently, post-translational modification of Ral has been shown to enhance the activities of RalGDS, indicating a functional role of this modification for transmitting its signal effectively (40).…”

K- and N-Ras Are Geranylgeranylated in Cells Treated with Farnesyl Protein Transferase Inhibitors

“…1, lanes 1-6). These proteins are believed to be farnesylated (21). Mevalonate incorporation into proteins that comprise a broad band between 21 and 28 kDa was not inhibited.…”