Cooperativity boosts affinity and specificity of proteins with multiple RNA-binding domains

Numerous cellular processes rely on the binding of proteins with high affinity to specific sets of RNAs. Yet most RNA-binding domains display low specificity and affinity in comparison to DNA-binding domains. The best binding motif is typically only enriched by less than a factor 10 in high-throughput RNA SELEX or RNA bind-n-seq measurements. Here, we provide insight into how cooperative binding of multiple domains in RNA-binding proteins (RBPs) can boost their effective affinity and specificity orders of magnitude higher than their individual domains. We present a thermodynamic model to calculate the effective binding affinity (avidity) for idealized, sequence-specific RBPs with any number of RBDs given the affinities of their isolated domains. For seven proteins in which affinities for individual domains have been measured, the model predictions are in good agreement with measurements. The model also explains how a two-fold difference in binding site density on RNA can increase protein occupancy 10-fold. It is therefore rationalized that local clusters of binding motifs are the physiological binding targets of multi-domain RBPs.

Automated summary

Most RNA-binding domains have lower specificity and affinity compared to DNA-binding domains. However, cooperative binding of multiple domains in RNA-binding proteins can significantly enhance their affinity and specificity. A thermodynamic model is presented to calculate the effective binding affinity of sequence-specific RBPs with any number of RBDs. The model predictions align well with measurements for seven proteins with known domain affinities. Furthermore, the model explains how a two-fold increase in binding site density on RNA can lead to a ten-fold increase in protein occupancy. Hence, it is proposed that local clusters of binding motifs serve as the physiological binding targets for multi-domain RBPs.