Reovirus μ1 structural rearrangements that mediate membrane penetration

Membrane penetration by nonenveloped reoviruses is mediated by the outer-capsid protein, mu 1 (76 kDa). Previous evidence has suggested that an autolytic cleavage in mu 1 allows the release of its N-terminally myristoylated peptide, mu 1N (4 kDa), which probably then interacts with the target-cell membrane. A substantial rearrangement of the remaining portion of mu 1, mu 1C (72 kDa), must also have occurred for mu 1N to be released, and some regions in mu 1C may make additional contacts with the membrane. We describe here a particle-free system to study conformational rearrangements of mu 1. We show that removal of the protector protein sigma 3 is not sufficient to trigger rearrangement of free mu 1 trimer and that free mu 1 trimer undergoes conformational changes similar to those of particle-associated mu 1 when induced by similar conditions. The mu 1 rearrangements require separation of the mu 1 trimer head domains but not the mu 1N/C autocleavage. We have also obtained a relatively homogeneous form of the structurally rearranged mu 1 (mu 1*) in solution. It is an elongated monomer and retains substantial alpha-helix content. We have identified a protease-resistant similar to 23-kDa fragment of mu 1*, which contains the largely alpha-helical regions designated domains I and II in the conformation of mu 1 prior to rearrangement. We propose that the mu 1 conformational changes preceding membrane penetration or disruption during cell entry involve (i) separation of the beta-barrell head domains in the mu 1 trimer, (ii) autolytic cleavage at the mu 1N/C junction, associated with partial unfolding of mu 1C and release of mu 1N, and (iii) refolding of the N-terminal helical domains of mu 1C, with which mu 1N was previously complexed, accompanied by dissociation of the mu 1 trimer.