Factors modulating filament formation by bovine glial fibrillary acidic protein, the intermediate filament component of astroglial cells.
Glial fibrillary acidic protein (GFAP) is soluble in low ionic strength solutions but shows a strong tendency toward assembly with increasing ionic strength as revealed by electron microscopy and turbidity measurements. Increasing K+, Na+, and Li+ concentrations cause an increase followed by a decrease in GFAP turbidity with a maximum at 200 mM, but their effects are much weaker than effects of divalent cations at the same ionic strength. Ca2+, Mg2+, Mn2+, and Ba2+ promote assembly at millimolar concentrations, and 10 microM Cu2+ causes rapid aggregation. The critical concentration for GFAP assembly was 0.08 +/- 0.04 mg/mL in 2 mM Tris-HCl, 60 mM KCl, and 1 mM CaCl2, pH 6.8. The Mr 38,000 rod domain of GFAP obtained by limited chymotryptic digestion is more soluble in 100 mM imidazole hydrochloride buffer, pH 6.8, than the intact molecule, and removal of the end pieces greatly reduces the ability of GFAP to form filaments. BNPS-skatole (2-[(2-nitrophenyl)sulfenyl]-3-methyl-3-bromoindolenine) treatment releases a Mr 30,000 N-terminus and a Mr 20,000 C-terminus. The Mr 30,000 polypeptide shows a higher affinity than the Mr 20,000 fragment for intact GFAP. Arginine and lysine at low concentrations slightly accelerate GFAP assembly, but above 100 mM both amino acids inhibit assembly. ATP, GTP, CTP, and UTP do not show significant effects on GFAP assembly. Dephosphorylation by alkaline phosphatase slightly reduces the assembly ability of GFAP, but phosphatase-treated GFAP still is assembly competent.
Yang
Z W
ZW
Chemistry of Behavior Program, Texas Christian University, Fort Worth 76129.
Babitch
J A
JA
D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D002412:Cations; D002417:Cattle; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism* / Q000648:ultrastructure; D066298:In Vitro Techniques; D008854:Microscopy, Electron; D008970:Molecular Weight; D009711:Nucleotides / Q000494:pharmacology; D010766:Phosphorylation; D011108:Polymers; D011487:Protein Conformation; D013116:Spinal Cord / Q000378:metabolism
D002412:Cations; D005904:Glial Fibrillary Acidic Protein; D009711:Nucleotides; D011108:Polymers