One of the proteins in the 0.5 M KCl eluate of rabbit reticulocyte polyribosomes which bind poly(A)-rich mRNA has been purified to apparent homogeneity using ammonium sulfate fractionation and phosphocellulose, hydroxylapatite, and diethylaminoethylcellulose column chromatography. The protein appears to contain two subunits of 66 700 and 56 400 apparent molecular weights with a 1:1 stoichiometry, since an apparent molecular weight of 110 000 was determined using Sephadex G-200 chromatography and an s020,w of 5.6 was obtained with rate-zonal sedimentation. The mRNA binding activity banded at pH 5.2-5.5 on isoelectric-focusing polyacrylamide gel electrophoresis. Protein-dependent binding appeared to be specific, since other natural or synthetic RNAs, including tRNA, ribosomal RNA, and poly(riboadenylic acid), were 90- to 250-fold less effective than mRNA at competing for binding of [3H]poly(adenylic acid)-rich mRNA. Poly(riboguanylic acid), however, was even more efficiently bound by this protein than mRNA.