The structures of multiple picornavirus polymerase
elongation complexes suggest that these enzymes use
a different molecular mechanism where translocation is
not strongly coupled to the opening of the active site
following catalysis. Here we present the 2.0- to
2.6-Å-resolution crystal structures and biochemical
data for 12 poliovirus polymerase mutants that together
show how proper enzyme functions and translocation
activity requires conformational flexibility of a loop
sequence in the palm domain B-motif. Within the loop,
the Ser288-Gly289-Cys290 sequence is shown to play
a major role in the catalytic cycle based on RNA
binding, processive elongation activity, and single
nucleotide incorporation assays. The structures show
that Ser288 forms a key hydrogen bond with Asp238,
the backbone flexibility of Gly289 is required for
translocation competency, and Cys290 modulates the
overall elongation activity of the enzyme. Some
conformations of the loop represent likely intermediates
on the way to forming the catalytically competent
closed active site, while others are consistent with a role
in promoting translocation of the nascent base pair out
of the active site. The loop structure and key residues
surrounding it are highly conserved, suggesting that the
structural dynamics we observe in poliovirus 3Dpol are
a common feature of viral RNA-dependent RNA
polymerases.
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