We have identified the Saccharomyces cerevisiae homolog of the signal recognition particle (SRP) and characterized its function in vivo. S. cerevisiae SRP is a 16S particle that includes a homolog of the signal sequence-binding protein subunit of SRP (SRP54p) and a small cytoplasmic RNA (scR1). Surprisingly, the genes encoding scR1 and SRP54p are not essential for growth, though SRP-deficient cells grow poorly, suggesting that SRP function can be partially by-passed in vivo. Protein translocation across the ER membrane is impaired in SRP-deficient cells, indicating that yeast SRP, like its mammalian counterpart, functions in this process. Unexpectedly, the degree of the translocation defect varies for different proteins. The ability of some proteins to be efficiently targeted in SRP-deficient cells may explain why previous genetic and biochemical analyses in yeast and bacteria did not reveal components of the SRP-dependent protein targeting pathway.