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J. Biol. Chem., Vol. 275, Issue 51, 40594-40600, December 22, 2000
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From the Department of Biochemistry, University of Iowa College of
Medicine, Iowa City, Iowa 52242
A major function of tropomyosin (TPM) in
nonmuscle cells may be stabilization of F-actin by binding
longitudinally along the actin filament axis. However, no clear
evidence exists in vitro that TPM can significantly affect
the critical concentration of actin. We previously made a
polymerization-defective mutant actin, GG (V266G, L267G). This actin
will not polymerize alone at 25 °C but will in the presence of
phalloidin or beryllium fluoride. With beryllium fluoride, but
not phalloidin, this polymerization rescue is cold-sensitive. We show
here that GG-actin polymerizability was restored by cardiac tropomyosin
and yeast TPM1 and TPM2 at 25 °C with rescue efficiency inversely
proportional to TPM length (TPM2 > TPM1 > cardiac
tropomyosin), indicating the importance of the ends in polymerization
rescue. In the presence of TPM, the apparent critical concentration of
actin is 5.5 µM, 10-15-fold higher than that of wild
type actin but well below that of the GG-actin alone (>20
µM). Non N-acetylated TPMs did not rescue GG-actin polymerization. The TPMs did not prevent cold-induced depolymerization of GG F-actin. TPM-dependent GG-actin
polymerization did not occur at temperatures below 20 °C.
Polymerization rescue may depend initially on the capture of unstable
GG-F-actin oligomers by the TPM, resulting in the strengthening of
actin monomer-monomer contacts along the filament axis.
Tropomyosin-dependent Filament Formation by a
Polymerization-defective Mutant Yeast Actin (V266G,L267G)*
*
This research is supported by a grant from the Muscular
Dystrophy Association of America (to P. A. R.), National Institutes of Health Grant GM-33689 (to P. A. R.), and a predoctoral fellowship from the Iowa Affiliate of the American Heart Association (to K. K. W.).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 all correspondence should be addressed. Tel.:
319-335-7911; Fax: 319-335-9570; E-mail:
peter-rubenstein@uiowa.edu.
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