Biology Reference
In-Depth Information
inactive (
A-b´
)(
B-a´
CCA) is expected to temporarily form a hexameric
intermediate ring. As reported previously, mutant pRNA
B-a´
CCA,
in which the C
18
C
19
A
20
bulge has been eliminated by insertion of
three bases 5
-UGG between A
99
and A
100
, is competent in hexameric
ring formation, but having no DNA packaging function.
150
If a five-
member pRNA ring were derived from a hexameric intermediate, one
pRNA should be dissociated from the intermediate hexameric ring to
form a five-member ring, as suggested by Rossmann
et al
.
111
However,
the “Six Minus One” set failed to produce infectious virions, which
means that “Six Minus One” cannot produce active five-member
pRNA rings.
As noted above, it is clearly revealed that five-member pRNA
rings derived from either a “Four Plus One” or a “Six Minus One”
approach were inactive in DNA packaging, arguing strongly against
the hypothesis that five pRNA make up a pentamer to drive the phi29
DNA-packaging motor (manuscript in preparation). Thus it is not
easy to account for how a pentameric pRNA ring could be formed
from an even-numbered binding unit (pRNA dimer) at a targeted
connector base that has even-numbered symmetry. This procedure
could potentially be applicable to any system in which the possible
stoichiometries need to be narrowed down and spurious options
eliminated.
′
Perspectives
This review covers mathematical approaches that have been applied
for stoichiometry quantification of the components of bacteriophage
phi29 assembly system. All studies led to the same conclusion that
phi29 DNA packaging motor contains six copies of pRNA, indicating
that these methods are feasible. However, the reliability of these
methods depends on the assay system. The more sensitive the assay
system, the more accurate; the lower the background noise, the
higher the reliability. For example, in phi29 the
in vitro
assembly sys-
tem is highly sensitive, since in the presence of all components, up to
10
9
infectious virions can be assembled, and not a single plaque is
detected if one of the components is omitted. The zero background
