The mechanism of RNA binding to TRAP: Initiation and cooperative interactions

The frp RNA-binding Attenuation Protein (TRAP) from Bacillus subtilis is an 11-subunit protein that binds a series of 11 GAG and UAG repeats separated by two to three-spacer nucleosides in trp leader mRNA. The structure of TR AP bound to an RNA containing 11 GAG repeats shows that the RNA wraps aroun d the outside of the protein ring with each GAG interacting with the protei n in nearly identical fashion. The only direct hydrogen bond interactions b etween the protein and the RNA backbone are to the 2'-hydroxyl groups on th e third G of each repeat. Replacing all 11 of these guanosines with deoxyri boguanosine eliminates measurable binding to TRAP. In contrast, a single ri boguanosine in an otherwise entirely DNA oligonucleotide dramatically stabi lizes TRAP binding, and facilitates the interaction of the remaining all-DN A portion with the protein. Studies of TRAP binding to RNAs with between 2 and 11 GAGs, UAGs, AAGs, or CAGs showed that the stability of a TRAP-RNA co mplex is not directly proportional to the number of repeats in the RNA. The se studies also showed that the effect of the identity of the residue in th e first position of the triplet, with regard to binding to TRAP, is depende nt on the number of repeats in the RNA. Together these data support a model in which TRAP binds to RNA by first forming an initial complex with a smal l subset of the repeats followed by a cooperative interaction with the rema ining triplets.