Molecular architecture of the vesicular stomatitis virus RNA polymerase.
Nonsegmented negative-strand (NNS) RNA viruses initiate infection by delivering into the host cell a highly specialized RNA synthesis machine comprising the genomic RNA completely encapsidated by the viral nucleocapsid protein and associated with the viral polymerase. The catalytic core of this protein-RNA complex is a 250-kDa multifunctional large (L) polymerase protein that contains enzymatic activities for nucleotide polymerization as well as for each step of mRNA cap formation. Working with vesicular stomatitis virus (VSV), a prototype of NNS RNA viruses, we used negative stain electron microscopy (EM) to obtain a molecular view of L, alone and in complex with the viral phosphoprotein (P) cofactor. EM analysis, combined with proteolytic digestion and deletion mapping, revealed the organization of L into a ring domain containing the RNA polymerase and an appendage of three globular domains containing the cap-forming activities. The capping enzyme maps to a globular domain, which is juxtaposed to the ring, and the cap methyltransferase maps to a more distal and flexibly connected globule. Upon P binding, L undergoes a significant rearrangement that may reflect an optimal positioning of its functional domains for transcription. The structural map of L provides new insights into the interrelationship of its various domains, and their rearrangement on P binding that is likely important for RNA synthesis. Because the arrangement of conserved regions involved in catalysis is homologous, the structural insights obtained for VSV L likely extend to all NNS RNA viruses.
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston MA 02115, USA.
Amal A
AA
Rahmeh
Andreas D
AD
Schenk
Eric I
EI
Danek
Philip J
PJ
Kranzusch
Bo
B
Liang
Thomas
T
Walz
Sean P J
SP
Whelan
D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D008958:Models, Molecular; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018116:Vesiculovirus / Q000201:enzymology*; D014764:Viral Proteins / Q000737:chemistry / Q000648:ultrastructure
D010750:Phosphoproteins; D014764:Viral Proteins; D012321:DNA-Directed RNA Polymerases