EFFECTS OF SUBTILISIN CLEAVAGE OF MONOMERIC ACTIN ON ITS NUCLEOTIDE-BINDING

Authors
Citation
A. Ooi et K. Mihashi, EFFECTS OF SUBTILISIN CLEAVAGE OF MONOMERIC ACTIN ON ITS NUCLEOTIDE-BINDING, Journal of Biochemistry, 120(6), 1996, pp. 1104-1110
Citations number
29
Categorie Soggetti
Biology
Journal title
ISSN journal
0021924X
Volume
120
Issue
6
Year of publication
1996
Pages
1104 - 1110
Database
ISI
SICI code
0021-924X(1996)120:6<1104:EOSCOM>2.0.ZU;2-Q
Abstract
The kinetics of ATP exchange on subtilisin-cleaved G-actin was investi gated by measuring the fluorescence of 1,N-6-ethenoadenosine 5'-tripho sphate. The apparent dissociation rate of ATP (k(-ATP)) was 2.8-fold l arger than that of intact G-actin in the presence of 300 mu M free Ca2 +. Analysis of the dependence of k(-ATP) on free Ca2+ showed that the dissociation rate constant of tightly bound Ca2+ was not significantly changed by subtilisin cleavage, On the other hand, an equilibrium bin ding study using 2-amino-5-methylphenoxy)methyl]-6-methoxyquinoline N, N,N',N'-tetraacetic acid (Quin 2) showed that the affinity of tightly bound Ca2+ for G-actin was reduced by about 13-fold after subtilisin t reatment. Consequently, the stabilization by Ca2+ of ATP was weak in c leaved G-actin. Furthermore, the kinetic analysis of ATP exchange reve aled that the binding equilibrium between ATP and divalent cat ion-fre e cleaved G-actin was much slower than that in the case of intact G-ac tin.