Protection of one of the two reactive thiol groups in F-actin by ATP and phalloidin

In F-actin, [A(SH)₅]_n, prepared from rabbit skeletal muscle, two thiol groups react with 2,4-dinitrophenyl-glutathionyl-disulfide, DNPSSG, to form [A(SH)₃(SSG)₂]_n. One of the two thiol groups reacts fast, (20 min), while the reaction of the second is slow (200 min). The fast reacting group has been identified as cysteine-373.In the presence of approximately one equivalent of ATP, only one of the thiol groups is reactive. The reaction product is [A(SH)₄SSG]_n. In comparison, the shielding effect of ADP is about 2 to 3 times smaller than that of ATP, while AMP is ineffective.The mushroom toxin phalloidin, which binds to polymeric actin, exhibits a similar protective activity as ATP and shields one thiol group from reaction with DNPSSG.We conclude from these data that in F-actin a second low affinity binding site for adenosin-nucleotides exists, which can be monitored by the reactivity of one of the two reactive thiol groups.