Interaction of gelsolin with covalently cross-linked actin dimer.
One of the two actin molecules in the ternary actin-gelsolin complex was selectively cross-linked to gelsolin when benzophenonemaleimide-actin (BPM-actin) was used [Doi, Y., Banba, M., & Vertut-Doi (1991a) Biochemistry 30, 5769-5777]. Here, we examine the interaction between gelsolin and BPM-actin dimer in which BPM-actin is covalently conjugated to unlabeled actin by p-phenylenedimaleimide (pPDM). BPM-actin dimer having an apparent molecular mass of 115 kDa is photo-cross-linked to gelsolin (90 kDa) more effectively than BPM-actin monomer in the presence of Ca2+, forming a cross-linked actin dimer-gelsolin (1:1) complex with a molecular mass of 210 kDa. The tight direct association of the dimer to gelsolin is shown by the titration of gelsolin with the fluorescently labeled dimer and by the higher concentration of phosphatidylinositol 4,5-bisphosphate required to inhibit the formation of BPM-dimer complex with gelsolin than that of BPM-monomer complex. However, an attempt to cross-link the two actin molecules in the ternary actin-gelsolin (2:1) complex by pPDM fails. The results argue that the topography of the two actin molecules in the actin-gelsolin (2:1) complex is similar, but not identical, to that of the barbed end of an actin filament.
Doi
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Laboratoire d'Enzymologie, CNRS, Gif Sur Yvette, France.
D009327:4-Chloro-7-nitrobenzofurazan; D000818:Animals; D001577:Benzophenones / Q000737:chemistry; D002118:Calcium / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D003432:Cross-Linking Reagents; D004533:Egtazic Acid / Q000494:pharmacology; D005456:Fluorescent Dyes; D018260:Gelsolin; D007700:Kinetics; D046911:Macromolecular Substances; D008301:Maleimides / Q000737:chemistry; D008840:Microfilament Proteins / Q000737:chemistry / Q000378:metabolism*; D008970:Molecular Weight; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols / Q000494:pharmacology; D010777:Photochemistry; D013552:Swine
D001577:Benzophenones; D002135:Calcium-Binding Proteins; D003432:Cross-Linking Reagents; D005456:Fluorescent Dyes; D018260:Gelsolin; D046911:Macromolecular Substances; D008301:Maleimides; D008840:Microfilament Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D010716:Phosphatidylinositols; D004533:Egtazic Acid; C046109:benzophenone-4-maleimide; C011328:N,N'-4-phenylenedimaleimide; D009327:4-Chloro-7-nitrobenzofurazan; D002118:Calcium