Correlation between Polymerizability and Conformation in Scallop β-Like Actin and Rabbit Skeletal Muscle α-Actin

In order to investigate the structural basis for functional differences among actin isoforms, we have compared the polymerization properties and conformations of scallop adductor muscle β-like actin and rabbit skeletal muscle α-actin. Polymerization of scallop Ca2+-actin was slower than that of skeletal muscle Ca2+-actin. Cleavage of the actin polypeptide chain between Gly-42 and Val-43 with Escherichia coli protease ECP 32 impaired the polymerization of scallop Mg2+-actin to a greater extent than skeletal muscle Mg2+-actin. When monomeric scallop and skeletal muscle Ca2+-actins were subjected to limited proteolysis with trypsin, subtilisin, or ECP 32, no differences in the conformation of actin subdomain 2 were detected. At the same time, local differences in the conformations of scallop and skeletal muscle actin subdomains 1 were revealed as intrinsic fluorescence differences. Replacement of tightly bound Ca2+ with Mg2+ resulted in more extensive proteolysis of segment 61–69 of scallop actin than in the case of skeletal muscle actin. Furthermore, segment 61–69 was more accessible to proteolysis with subtilisin in polymerized scallop Ca2+-actin than in polymerized skeletal muscle Ca2+-actin, indicating that, in the polymeric form, the nucleotide-containing cleft is in a more open conformation in β-like scallop actin than in skeletal muscle α-actin. We suggest that this difference between scallop and skeletal muscle actins is due to a less efficient shift of scallop actin subdomain 2 to the position it has in the polymer. The possible consequences of amino acid substitutions in actin subdomain 1 in the allosteric regulation of the actin cleft, and hence in the different stabilities of polymers formed by different actins, are discussed.