Purification and properties of virus particles, infectious subviral particles, cores and VP7 crystals of African horsesickness virus serotype 9.
Methods were developed for the purification, at high yield, of four different particle types of African horsesickness virus serotype 9 (AHSV-9). These products included virus particles purified on CsCl gradients which contain proteins apparently directly comparable to those of bluetongue virus (VP1 to VP7); virus particles purified on sucrose gradients which also contain, as a variable component, protein NS2; infectious subviral particles (ISVPs), containing chymotrypsin cleavage products of VP2; and cores, obtained by treating purified ISVPs with 1 M-MgCl2 to remove the components of the outer capsid layer (VP5 and VP2 cleavage products). Additional protein bands migrating with apparent M(r)s lower than that of VP5 were detected during SDS-PAGE analysis of virus particles. These appear to be conformational variants of VP5 and are identified as VP5' and VP5". BHK-21 cells infected with this strain of AHSV-9 produce large quantities of flat, usually hexagonal crystals of VP7, a major group antigen and core protein; these were also purified. Either 20 mg of virus particles, 20 mg of ISVPs or 10 mg of cores as well as 20 mg of VP7 crystals could be purified from approximately 8 x 10(9) infected cells. None of the preparations of particles or crystals showed any detectable contamination with BHK-21 cell proteins or antigens, as determined by SDS-PAGE or indirect ELISA. Virus particle and ISVP preparations had similar specific infectivities for BHK-21 cells (approximately 1 x 10(9) TCID50/A260 unit) but the infectivity of cores was approximately 10(5)-fold lower.
Burroughs
J N
JN
AFRC Institute for Animal Health, Pirbright Laboratory, Woking, Surrey, U.K.
O'Hara
R S
RS
Smale
C J
CJ
Hamblin
C
C
Walton
A
A
Armstrong
R
R
Mertens
P P
PP
D000356:African Horse Sickness Virus / Q000145:classification / Q000302:isolation & purification* / Q000472:pathogenicity / Q000648:ultrastructure; D000818:Animals; D000956:Antigens, Viral / Q000302:isolation & purification* / Q000648:ultrastructure; D002478:Cells, Cultured; D006224:Cricetinae; D003460:Crystallization; D008854:Microscopy, Electron; D014758:Viral Core Proteins / Q000302:isolation & purification* / Q000648:ultrastructure; D014774:Virulence
D000956:Antigens, Viral; D014758:Viral Core Proteins; C069264:VP7 major core antigen, African horsesickness virus