Purification, crystallization, and preliminary X-ray analysis of L-A: a dsRNA yeast virus.
TheL-A virus (LAV) particle is a specialized compartment for the transcription and replication of double-stranded RNA. It is 390 A in diameter and infects yeast. The particle is formed by a capsid containing 120 copies of a 680-residue gene product arranged with T = 1 icosahedral symmetry, approximately two copies of an RNA-directed RNA polymerase, and a 4.6-kb linear, duplex RNA. LAV crystals diffracting to at least 4.5-A resolution were grown in a combination of polyethylene glycol 8000, ethylene glycol, and lithium chloride. Following crystallization the reservoir solution was replaced by a 2x concentrated reservoir solution in order for ethylene glycol to function as a cryoprotectant even though initial crystals would not grow at sufficiently high concentrations of ethylene glycol for cryoprotection. A complete data set was collected to 6-A resolution from a frozen crystal obtained with this procedure. The crystals belong to space group P2(1). The unit cell dimensions are a = 406.7 A, b = 403.3 A, c = 572.5 A, beta = 90.3 degrees with two virus particles in the unit cell. The particle orientation was determined with the rotation function and the particle center was estimated on the basis of packing considerations.
Naitow
H
H
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Canady
M A
MA
Lin
T
T
Wickner
R B
RB
Johnson
J E
JE
D002213:Capsid / Q000737:chemistry; D003460:Crystallization; D018628:Gene Dosage; D015683:Gene Products, gag / Q000235:genetics; D015684:Gene Products, pol / Q000235:genetics; D008970:Molecular Weight; D012330:RNA, Double-Stranded / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000821:virology*; D019197:Totivirus / Q000737:chemistry*; D014771:Virion / Q000737:chemistry* / Q000302:isolation & purification*; D014961:X-Ray Diffraction / Q000379:methods
D015683:Gene Products, gag; D015684:Gene Products, pol; D012330:RNA, Double-Stranded; D012324:RNA-Dependent RNA Polymerase