Prenylation is a posttranslational modification having a critical role in the proper biological function of many proteins. This modification involves the attachment of a 15- or 20- carbon isoprenoid group to a cysteine residue near the C-terminus of the target substrate by protein farnesyltransferase (FTase) or protein geranylgeranyltransferase type I (GGTase-I), respectively. Following prenylation, many prenylated proteins undergo subsequent processing steps that assist in localization of the prenylated protein to the cell membrane and/or formation of protein-protein interactions essential for biological function. A long-held model for substrate selectivity involves both protein prenyltransferases recognizing their respective substrates based on the presence of a C-terminal “Ca1a2X” sequence, where C denotes the cysteine residue to be modified and a1, a2 and X residues provide for sequence selectivity leading to modification by FTase and/or GGTase-I. Recent studies in yeast, however, suggest the potential for FTase recognition and modification of sequences that diverge from the canonical “Ca1a2X” sequence such as the recently reported five amino acid “C(x)3X” motif for FTase substrates. Looking towards additional unexplored options for sequence motifs that can undergo prenylation, we explored the ability of mammalian FTase and GGTase-I to recognize, bind, and modify three amino acid “Cxx” sequences, focusing on ones derived from both yeast screening experiments and analysis of the human proteome. In assays using purified enzyme, we observed evidence for the sequence-selective modification of multiple Cxx sequences. Steady-state kinetic analyses of these peptides indicate that some sequences exhibit sufficient reactivity to be potentially modified within the cell by endogenous prenyltransferase activity. These findings expand the substrate pool of sequences that can be potentially prenylated, further refine our understanding of substrate recognition by FTase and GGTase-I, and suggest the possibility of a new prenylated protein class within the human proteome.

Expanding the potential prenylome through unanticipated prenylation of non-canonical C-terminal peptide sequences

256th National Meeting (2018)

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