Arch Virol (2005) 150: 2037–2050
Characterization of the nucleic acid binding activity
of inner core protein VP6 of African horse sickness virus
P. J. de Waal and H. Huismans
Department of Genetics, University of Pretoria, Pretoria, South Africa
Received December 16, 2004; accepted April 4, 2005
Published online June 28, 2005
Summary. Minor structural protein VP6 is the putative helicase of African
horse sickness virus (AHSV), of the genus Orbivirus in the Reoviridae family.
We investigated how the protein interacts with double-stranded (ds) RNA and
other nucleic acids. Binding was assayed using an electrophoretic migration
retardation assay and a nucleic acid overlay protein blot assay. VP6 bound double
and single stranded RNA and DNA in a NaCl concentration sensitive reaction.
Of six truncated VP6 peptides investigated, two partially overlapping peptides
were found to bind dsRNA at pH 7.0, while other peptides with the same overlap
did not. The distinction between the peptides appeared to be the pI which ranged
from more than 8.0 to just above 6.0. Changing the pH of the binding buffer
modiﬁed the binding activity. Regardless of assay conditions, only peptides with
a speciﬁc region of amino acids in common, showed evidence of binding activity.
No sequence homology was identiﬁed with other binding domains, however, the
presence of charged amino acids are assumed to be important for binding activity.
The results suggested dsRNA binding in the blot assay was strongly affected by
the net charge on the peptide.
African horse sickness virus (AHSV) is the causative agent of African horse sick-
ness (AHS), an infectious, but non-contagious vector-borne disease of equines.
The disease is characterized by a very high mortality rate of up to 95% in naive
populations of horses  and it is endemic in South Africa and many other parts
of sub-Saharan Africa. Sporadic outbreaks have occurred in a variety of countries
ranging from Egypt, the Middle East, South West Asia, India, Cyprus and Spain
[15, 16]. The AHSV virion has a double-stranded (ds) RNA genome  and
belongs to the genus Orbivirus, in the family Reoviridae . The AHSV particle
consists of an outer capsid containing two major structural proteins VP2 and VP5