The CCR4 family proteins are 3'-5'-deadenylases that function in the first step 
of the degradation of poly(A) mRNA. Here we report the purification to 
homogeneity of the yeast CCR4 protein and the analysis of its substrate 
specificities. CCR4 deadenylated a 7N+23A substrate (seven nucleotides followed 
by 23 A residues) in a distributive manner. Only small differences in CCR4 
activity for different A length substrates were observed until only 1 A residue 
remained. Correspondingly, the K(m) for a 25N+2A substrate was found to be at 
least 20-fold lower than that for a 26N+1A substrate, although their V(max) 
values differed by only 2-fold. In addition, the total length of the RNA was 
found to contribute to CCR4 activity: up to 17 nucleotides (not necessarily 
poly(A)) could be recognized by CCR4. Poly(U), poly(C), and poly(G) were also 
found to be 12-30-fold better inhibitors of CCR4 compared with poly(A), 
supporting the observation that CCR4 contains a non-poly(A)-specific binding 
site. Surprisingly, even longer substrates (>/=45 nucleotides) stimulated CCR4 
to become a processive enzyme, suggesting that CCR4 undergoes an additional 
transition in the presence of such substrates. CCR4 also displayed no difference 
in its activity with capped or uncapped RNA substrates. These results indicate 
that CCR4 recognition of its RNA substrates involves several features of the RNA 
that could be sites in vivo for controlling the rate of specific mRNA 
deadenylation.