ANALYSIS OF LONG-RANGE STRUCTURAL EFFECTS INDUCED BY DNASE-I INTERACTION WITH ACTIN MONOMERIC FORM OR COMPLEXED TO CAPZ

Two fundamental properties of monomeric actin were examined in this st udy, ie its interaction with DNase-I, and the inhibition of endonuclea se activity consecutive to the association of the two molecules. In pa rticular, the topological independence between catalytic site of DNase -I and interface with actin, structural changes in actin monomer and t he absence of conformational changes in DNase-I were described. We dem onstrated a loss of flexibility of antigenic structures in actin subdo main I (ie epitopes 18-28 and 95-105) as well as modification in the e xposure of Cys10 and Cys374 after DNase-I binding. Furthermore, the co nformational changes induced by DNase-I into the actin molecule weaken ed the interaction of CapZ to its binding site located in the C-termin al region of actin monomer. These structural changes were time-depende nt. When actin was cleaved in the DNase-I binding loop (sequence 38-52 ) at position 42 by E coli A2 strain protease, a tight DNase-I binding to split actin and the conformational changes were still observed, wh ereas the DNase-I inhibition activity was completely abolished. Finall y, when we substitute Ca2+ by Mg2+ (ATP-Mg2+ monomeric actin) which in duces a tighter conformation of actin and partially restores the inhib itory ability of split actin, long-range conformational effects of DNa se-I are prevented and the ternary complex DNase-I-actin-CapZ is obtai ned.