Biochemical and theoretical approach to localization of metal-ion-binding sites in the actin primary structure.
The number of Ca2+ ions bound at sites other than the single high-affinity site in CaCl2-induced polymers of rabbit skeletal muscle, chicken gizzard, and bovine aorta actin was determined. The polymer of skeletal muscle and aorta actin contained 4 mol Ca2+/mol, whereas gizzard actin only 3 mol weakly bound Ca2+/mol monomer. This difference correlates with the deletion in smooth muscle gamma-actin of one out of four NH2-terminal acidic residues typical of skeletal and smooth muscle alpha-actin isoforms, suggesting that this additional acidic residue in alpha-actins is involved in the weak binding of cations which is essential for polymerization. This experimental result, as well as a theoretical analysis of the actin primary structure, argue against the implication of the NH2-terminal acidic residues in the high-affinity site for divalent cation. The analysis of the actin primary structure aimed at identification of sequences resembling the known Ca2+-binding patterns has revealed the absence of an EF-hand Ca2+-binding site. The best match was obtained between the sequence of the 292-301 segment and that of Ca2+ site in lectins. However, in the light of experimental data discussed, it is more plausible that the actual high-affinity Ca2+ site in actin involves sequentially distant residues from the NH2- and COOH-terminal portions of the polypeptide chain.
Strzelecka-Goøaszewska
H
H
Department of Muscle Biochemistry, Nencki Institute of Experimental Biology, Warsaw, Poland.
Boguta
G
G
Zmorzyński
S
S
Moraczewska
J
J
D000199:Actins / Q000032:analysis*; D000345:Affinity Labels; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000032:analysis*; D002417:Cattle; D002645:Chickens; D005895:Gizzard, Avian / Q000032:analysis; D009131:Muscle, Smooth, Vascular / Q000032:analysis*; D009132:Muscles / Q000032:analysis*; D011108:Polymers / Q000032:analysis; D011189:Potassium Chloride / Q000494:pharmacology; D011817:Rabbits; D013050:Spectrometry, Fluorescence
D000199:Actins; D000345:Affinity Labels; D011108:Polymers; D011189:Potassium Chloride; D002118:Calcium