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Journal of Virology, February 1999, p. 1108-1117, Vol. 73, No. 2
0022-538X/99/$00.00+0
Envelope Formation Is Blocked by Mutation of a
Sequence Related to the HKD Phospholipid Metabolism Motif in the
Vaccinia Virus F13L Protein
Rachel L.
Roper and
Bernard
Moss*
Laboratory of Viral Diseases, National
Institute of Allergy and Infectious Diseases, Bethesda, Maryland
20892-0445
Received 22 July 1998/Accepted 20 October 1998
The outer envelope of the extracellular form of vaccinia virus is
derived from Golgi membranes that have been modified by the insertion
of specific viral proteins, of which the major component is the 37-kDa,
palmitylated, nonglycosylated product of the F13L gene. The F13L
protein contains a variant of the HKD (His-Lys-Asp) motif, which is
conserved in numerous enzymes of phospholipid metabolism. Vaccinia
virus mutants with a conservative substitution of either the K (K314R)
or the D (D319E) residue of the F13L protein formed only tiny plaques
similar to those produced by an F13L deletion mutant, were unable to
produce extracellular enveloped virions, and failed to mediate
low-pH-induced fusion of infected cells. Membrane-wrapped forms of
intracellular virus were rarely detected in electron microscopic images
of cells infected with either of the mutants. Western blotting and
pulse-chase experiments demonstrated that the D319E protein was less
stable than either the K314R or wild-type F13L protein. Most striking,
however, was the failure of either of the two mutated proteins to
concentrate in the Golgi compartment. Palmitylation, oleation, and
partitioning of the F13L protein in Triton X-114 detergent were
unaffected by the K314R substitution. These results indicated that the
F13L protein must retain the K314 and D319 for it to localize in the Golgi compartment and function in membrane envelopment of vaccinia virus.
*
Corresponding author. Mailing address: Laboratory of
Viral Diseases, National Institute of Allergy and Infectious Diseases, Building 4, Room 229, 4 Center Dr. MSC 0445, National Institutes of
Health, Bethesda, MD 20892-0445. Phone: (301) 496-9869. Fax: (301)
480-1147. E-mail: bmoss{at}nih.gov.
Journal of Virology, February 1999, p. 1108-1117, Vol. 73, No. 2
0022-538X/99/$00.00+0
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