S-100 protein, but not calmodulin, binds to the glial fibrillary acidic protein and inhibits its polymerization in a Ca(2+)-dependent manner.
S-100 protein, a Ca(2+)-binding protein of the EF-hand type, interacts with the glial fibrillary acidic protein (GFAP) in a Ca(2+)-dependent manner. The binding of S-100 protein to GFAP was investigated by fluorescence spectroscopy using acrylodan-S-100 protein and cross-linking experiments using the bifunctional cross-linker, disuccinimidyl suberate. The binding affinity was observed to be in the nanomolar range with a stoichiometry of 2 mol of GFAP/mol of S-100 protein (dimer). S-100 protein was found to inhibit the polymerization of GFAP in a dose- and Ca(2+)-dependent manner, with a half-maximal effect at an S-100 protein/GFAP molar ratio of 0.2 and maximal effect at a molar ratio of 0.5. Identical results were obtained irrespective of whether the unfractionated bovine brain S-100 protein mixture (S-100a plus S-100b), S-100ao, S-100a, or S-100b was used. S-100 protein was observed to be maximally effective as an inhibitor of GFAP polymerization at approximately 3 microM free Ca2+. Calmodulin neither bound to GFAP nor inhibited its polymerization. Altogether, the present results suggest that S-100 protein might be involved in the regulation of the state of assembly of glial filaments by binding to and sequestering unpolymerized GFAP.
Bianchi
R
R
Department of Experimental Medicine and Biochemical Sciences, University of Perugia, Italy.
Giambanco
I
I
Donato
R
R
D015081:2-Naphthylamine / Q000031:analogs & derivatives; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000302:isolation & purification / Q000378:metabolism*; D002417:Cattle; D002852:Chromatography, Ion Exchange; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D005456:Fluorescent Dyes; D005904:Glial Fibrillary Acidic Protein / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D011485:Protein Binding; D009418:S100 Proteins / Q000302:isolation & purification / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013388:Succinimides
D002147:Calmodulin; D003432:Cross-Linking Reagents; D005456:Fluorescent Dyes; D005904:Glial Fibrillary Acidic Protein; D046911:Macromolecular Substances; D009418:S100 Proteins; D013388:Succinimides; C038518:acrylodan; D015081:2-Naphthylamine; D002118:Calcium; C019358:disuccinimidyl suberate