Sbp1 is a single-stranded RNA-binding protein that contains two RNA recognition motifs and an arginine–glycine–glycine rich domain. In this study, we investigated molecular interactions important for the transcript-specific translational regulation of Sbp1 on an essential mRNA that encodes the poly(A)-binding protein Pab1 in vitro. Our results demonstrate that the RNA-binding property and RGG domain of Sbp1 are important for its function.