title: Studies on the antigenic sites of actin: a comparative study of the immunogenic crossreactivity of invertebrate actins.
abstract: The structural homologies of invertebrate actins with cytoplasmic vertebrate actins have recently been substantiated by comparative sequence analyses. This suggests that cytoplasmic actin is the ancestral precursor of smooth and striated muscle actin in vertebrates. We have raised antibodies in rabbits against a number of invertebrate muscle actins and have characterized the antisera by means of the highly sensitive ELISA method, which allows quantitation of nanomolar amounts of actin. Despite the fact that the invertebrate actins examined are very similar in primary structure, our results indicate that antibodies raised against them clearly distinguish between only a few amino-acid substitutions, and that the immunoreactivities quantitatively reflect the genetic divergence of this ubiquitous conservative protein. Examination of several proteolytic fragments of scallop actin for immunoreactivity with the homologous antiserum suggests that the major antigenic sites of actin are located within the amino terminal region of the molecule, while a carboxy terminal fragment comprising residues 69-372 exhibits very weak crossreactivity. Immunoadsorption experiments further indicate that species-specific antibodies are directed to antigenic determinants in the N-terminal region. This finding is supported by an examination of the effects of chemical modifications to Tyr, His, Arg, and Cys residues on the immunoreactivity of actin. Interaction with DNAase I markedly decreases the immunoreactivity of actin. This is consonant with the finding that the amino terminal peptide comprising residues 1-207 inhibits DNAase I, whilst a tryptic fragment fails to bind to the enzyme. The interaction is abolished by EDTA and the removal of the tightly bound cation is accompanied by a conformational change, shown by shifts in circular dichroic spectra. The possible involvement of the amino terminal peptide of actin in cation binding is discussed.
mesh_terms: D000199:Actins / Q000276:immunology*; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D000906:Antibodies / Q000276:immunology; D001665:Binding Sites; D003429:Cross Reactions; D003850:Deoxyribonuclease I; D004706:Endodeoxyribonucleases / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis*; D007448:Invertebrates; D010446:Peptide Fragments / Q000276:immunology
chemical_list: D000199:Actins; D000596:Amino Acids; D000906:Antibodies; D000939:Epitopes; D010446:Peptide Fragments; D004706:Endodeoxyribonucleases; D003850:Deoxyribonuclease I