Equilibrium studies on the association of the nuclear poly(A) binding protein with poly(A) of different lengths

The nuclear poly(A) binding protein (PABPN1) binds the growing poly(A) tail during pre-mRNA 3'-end processing, stimulating its elongation and controlling its final length. Here we report binding studies of PABPN1 to poly(A) in solution. Quantitative fluorescence titration was used to determine the stoichiometry, intrinsic affinity, and cooperativity of binding to a series of size-fractionated poly(A). The intrinsic association constant K-i was about 2 x 10(6) M-1 for oligo(A) and all size classes of poly(A). The binding of PABPN1 to poly(A) was enhanced by protein-protein interactions which were, however, weak (cooperativity parameter omega < 50). No significant change of cooperativity could be detected with increasing polynucleotide length in the range of 140-450 nucleotides. An average binding site size n of 11-14 was found for all poly(A) lengths, which is close to the minimal site size in found for binding to oligo(A). The data are discussed with respect to the previous observation of two different forms of the poly(A)-PABPN1 complex.