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Virology 2015
RNA Virus RdRP Enzymes
General Comments
 An
RNA virus does not view replication of
the genome and expression of the
genome as separate events.
 An RNA virus is an RNA replicon in a
world full of DNA replicons.
 For most RNA viruses, everything takes
place in the same compartment.
 A biological arms race has shaped the
evolution of RNA viruses
Issues, problems and strategies related to the
production of many copies of progeny viral
RNA (replication)
The nature of the template (pp 170-171)
secondary structure
nucleocapsids
The nature of the enzymes (pp 171-175)
only viruses have RdRP (almost)
The nature of the reactions (pp 175-182)
initiation and priming
elongation/unwinding
balancing + and – strand synthesis
The Nature of the Template
RNA secondary structure is
complex
Secondary structures help
regulate RNA function
Shape is usually the key
Replication machinery must
deal with the shape
Example: CSE4 of alphaviruses
Alphaviruses are members of a genus of the
Togavirus family
SS + RNA, T=4, envelope, wide host range
Replication initiates at 3’ CSE using
non-structural virus proteins and host factors
Shown for salmonid alphavirus 3 (SAV3)
Example for
nucleocapsids:
VSV
Vesicular Stomatitis
Virus: vesiculovirus
genus of
Rhabdoviridae
(best-studied
rhabdovirus)
Rhabdoviridae are – ssRNA with a
“bullet” shape
enveloped
Wide host range among
rhabdoviridae
Example: VSV Replication requiresor RNP
nucleocapsid
“Transcription” of genomic RNA produces + sense
messages and viral proteins
Production of full-length + or – RNA requires
coated template
The Nature of the Enzymes
RdRp-only viruses have it???
Increasing understanding of function of RNA
in an uninfected cell
Common domains or “motifs”
RdRp Motifs:
A: nucleotide recognition/binding, phosphoryl transfer: “two-metal” mechanism
B: nucleotide recognition/binding
C: active site, phosphoryl transfer: “two-metal” mechanism
D: structure and shape of enzyme
E: primer binding
F: nucleotide entry tunnel (not shown above)
(A, B, C, D in all pols)
Common shape
Polio 3Dpol (above), HCV
(below)
Shaped like a right hand
Key features: Palm,
Thumb, Fingers
Closed configuration
Generic shape-no two are
identical
Shape changes affect
biological properties
Location of Domains
The conserved structural polymerase
motifs. (a–c) The polymerase core of IBDV
VP1 (a) compared with the equivalent cores
in FMDV (b) and bacteriophage φ6 (c)
RDRPs. The secondary structural elements
containing the conserved motifs are colored
as follows: A, red; B, green; C, yellow; D,
purple; E, orange; F, blue. The N- and Cterminal domains are shown as thin ribbons.
FMDV = Foot and mouth disease virus
IBDV = infectious bursal disease virus
RdRp with template
Below: Polio Rdrp with RNA in substrate
channel, top removed
Right (A) WNV Rdrp with motifs and
position of substrate, template and
product
(B) 90 degree rotation of A
The Nature of the Reactions:
Initiation and Priming
de novo Initiation
Prime and realign
Proposed mechanism for:
arenaviridae, bunyaviridae:
ss ambisense RNA
segmented genome
enveloped
wide host range-rodents
arena (Machupo)
bunya (hantaviruses)
De novo refers to lack of requirement for 3’ OH
Primer Dependent Initiation
Protein priming
VPg of picornaviruses = best example
ss + RNA, AAAAA, icosahedral,
many species
22 aa covalently linked
Virion RNA
Removed upon infection to produce
viral mRNA
Polio Gene Products
 3AB
= membrane anchor
 3B = VPg
 3C or 3CD = protease
 3D = polymerase
Replication machine anchored to membrane
Addition of U to 3AB
Coupled to cleavage of 3B from 3AB
to produce 3A and 3B=VPg-pUpU
Uridylylated VPg transferred to 3’ end
5’ VPg-pUpU 
3’
ApApApApNNNNN 5’
Host proteins
PCBP
And
PABP
Are
Important
The Nature of the Reactions:
Unwinding
Helicase activity
Example:
Flaviviruses: Yellow Fever Virus,
West Nile Virus,
Hepatitis C Virus
RNA helicase of flavivirus a member
of helicase superfamily 2
Helicase a target for therapy?
ATP dependent
Mechanism?
See p 180
Not all helicases function in this way
Phage phi 6 is a dsRNA phage
whose polymerase/helicase
works differently
The Nature of the Reactions:
Balancing + and - Synthesis
+/- shift not well-characterized
Diverse mechanisms?
Differential stability of + and -?
Differential synthesis of + and -?
Deterioration of cells/buildup of virus factories
affects stoichiometry?