The Cys3-His1 Zinc binding motif of the hRSV M2-1 tetramer modulates its dissociation to folded apo-monomers.

SEBASTIÁN ESPERANTE, MARIA GABRIELA NOVAL, TAMARA ANTONELLA, AND GONZALO DE PRAT GAY.

Granada

Congreso; 15th International Conference on Negative Strand Virus Meeting; 2013

The M2-1 protein of human respiratory syncytial virus (hRSV) functions as a transcription antiterminator by increasing polymerase processivity, thus enhancing readthrough of intergenic junctions. It is present only in pneumoviruses, namely hRSV and metapneumovirus. The hRSV M2-1 is a tight tetramer bearing a Cys3-His1 motif present in eukariotic transcription factors, which binds one zinc atom per monomer, and was shown to be essential for protein function by mutational analysis of the zinc coordinating residues. Using chemical perturbation, cross-linking, size exclusion chromatography, and dynamic light scattering, we showed that removal of the zinc atom leads to an Apo-M2-1 monomer. However, the secondary structure and stability of the apo-monomer is indistinguishable from the M2-1 holo-tetramer, indicating conservation of the native fold in the monomer. Thus, the complete exposure of its unique trp residue is not indicative of tertiary structure loss but is a sensitive reporter for metal removal and quaternary structure. Dissociation is much increased at pH 5.0 compared to pH 7.0 which srongly suggests that pH modulates zinc removal by histidine protonation and this, in turn, dissociation. Although no specific RNA sequence has been identified so far for M2-1, the monomeric apo form binds tRNA at least as well as the holo-tetramer. On the other hand, the monomer appears not to be competent for interacting with the phosphpoprotein P, the essential RNA polymerase cofactor. We discuss the results in connection with possible consequences in hRSV RNA transcription and genome replication.