Endoplasmic reticulum membrane localization of Rce1p and Ste24p, yeast proteases involved in carboxyl-terminal CAAX protein processing and amino-terminal a-factor cleavage

• DOI: 10.1073/pnas.95.19.11175
• Publication Date: 2002-07-26T14:42:40Z
• Authors: Walter K. Schmidt; Amy Tam; Konomi Fujimura-K...; Susan Michaelis
• Keywords: 0303 health sciences; Saccharomyces cerevisiae Proteins; Genetic Complementation Test; Fluorescent Antibody Technique; Membrane Proteins; Metalloendopeptidases; Saccharomyces cerevisiae; Cell Fractionation; Endoplasmic Reticulum; Fungal Proteins; 03 medical and health sciences; Endopeptidases; Proprotein Convertases; Protein Processing, Post-Translational

Proteins terminating in the CAAX motif, for example Ras and yeast a-factor mating pheromone, are prenylated, trimmed of their last three amino acids, carboxyl-methylated. The enzymes that mediate these activities, collectively referred to as processing components, have been identified genetically Saccharomyces cerevisiae . Whereas Ram1p/Ram2p prenyltransferase is a cytosolic soluble enzyme, sequence analysis predicts other Rce1p Ste24p proteases Ste14p methyltransferase, contain multiple membrane spans. To determine intracellular site(s) at which occurs, we examined localization by subcellular fractionation indirect immunofluorescence. We find both associated with endoplasmic reticulum (ER) membrane. In addition having role processing, protease catalyzes first two cleavage steps remove amino-terminal extension from precursor, suggesting step maturation also occurs ER consistent presence carboxyl-terminal dilysine retrieval although mutation this motif does not result mislocalization Ste24p. Because ultimate destination or most proteins, our results imply mechanism must exist routing proteins cellular sites.