The respiratory syncytial virus transcription antiterminator M2-1 is a highly stable, zinc binding tetramer with strong pH dependent dissociation and a monomeric unfolding intermediate

ESPERANTE, S.; CHEMES, L.B.; SÁNCHEZ, I.E.; DE PRAT-GAY, G.

BIOCHEMISTRY

AMER CHEMICAL SOC

Año: 2011 vol. 50 p. 8529 - 8539

0006-2960

The human respiratory syncytial virus M2-1 transcription antiterminator is an essential elongation factor required by the RNA polymerase for effective transcription beyond the first two non-structural genes. Its exclusive presence in pneumovirus among all paramyxovirus, suggests a unique function within this small genus. With the aim of understanding its biochemical properties we investigated this α-helical tetramer by making use of a biophysical approach. We found that the tetramer hydrodynamic radius is considerably extended at high ionic strength, and measured its zinc content to be one atom per monomer. Dissociation-unfolding experiments show a fully reversible and concentration dependent cooperative transition, but secondary and tertiary structure changes are uncoupled at lower protein concentrations. We detect the presence of a monomeric intermediate, which can be classified as a “late molten globule” with substantial secondary and tertiary structure. Global fittings of experiments from three different probes at two M2-1 concentrations provide a free energy of dissociation-unfolding of - 36.8 ± 0.1 kcal mol-1, corresponding to a tight dissociation constant of 10-27 M3 at pH 7.0. The tetramer affinity is strongly governed by pH, with a free energy change of 12 kcal mol-1 on going from pH 7.0 to pH 5.0 (KD = 10-16 M3). The drastic changes that take place within a pH range compatible with cellular environment strongly suggests a regulatory effect of pH on M2-1 structure and biochemical properties, likely affecting transcription and interaction with proteins and RNA