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J. Biol. Chem., Vol. 275, Issue 23, 17510-17516, June 9, 2000
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From the Six RNA (pRNA) molecules form a hexamer, via
hand-in-hand interaction, to gear bacterial virus phi29 DNA
translocation machinery. Here we report the pathway and the conditions
for the hexamer formation. Stable pRNA dimers and trimers were
assembled in solution, isolated from native gels, and separated by
sedimentation, providing a model system for the study of RNA dimers and
trimers in a protein-free environment. Cryo-atomic force microscopy
revealed that monomers displayed a
A Dimer as a Building Block in Assembling RNA
A HEXAMER THAT GEARS BACTERIAL VIRUS phi29 DNA-TRANSLOCATING
MACHINERY*
§,
Department of Pathobiology, Purdue
University, West Lafayette, Indiana 47907 and the ¶ Department of
Molecular Physiology and Biological Physics, University of Virginia,
Charlottesville, Virginia 22908
outline, dimers exhibited an
elongated shape, and trimers formed a triangle. Dimerization of pRNA
was promoted by a variety of cations including spermidine, whereas
procapsid binding and DNA packaging required specific divalent cations,
including Mg2+, Ca2+, and
Mn2+. Both the tandem and fused pRNA dimers with
complementary loops designed to form even-numbered rings were active in
DNA packaging, whereas those without complementary loops were inactive.
We conclude that dimers are the building blocks of the hexamer, and the
pathway of building a hexamer is: dimer 
tetramer hexamer. The
Hill coefficient of 2.5 suggests that there are three binding sites with cooperative binding on the surface of the procapsid. The two
interacting loops played a key role in recruiting the incoming dimer,
whereas the procapsid served as the foundation for hexamer assembly.
*
This work was supported by National Science Foundation
Grants MCB9723923 (to P. G.) and DBI9730060 (to Z. S.) and National Institutes of Health Grants GM59944 (to P. G.) and RRO7720 (to Z. S.)
and by the Integrated Biotechnology Corp. (to P. G.).The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
To whom correspondence should be addressed: Purdue Cancer
Center, B-36 Hansen Life Science Research Bldg., Purdue University, West Lafayette, IN 47907. Tel.: 765-494-7561; Fax: 765-496-1795; E-mail: guo@vet.purdue.edu.
Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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