Changes in the cooperativity of diaphragm-associated acetylcholinesterase induced by monovalent (Na+, Li+) cations.
Acetylcholinesterase (AChE) activity has been determined using homogenized rat diaphragm and soluble AChE from the eel Electrophorus electricus, using as a substrate different amounts of acetylthiocholine in the presence or absence of 115 mM NaCl or LiCl. With LiCl the KM values derived from Lineweaver-Burk plots are found to be 470-650 and 1045-1425 microM without cations or with NaCl. The cooperativity of the enzyme is increased when cations are added to the homogenate, as demonstrated by changes of the Hill coefficient. With soluble AChE, only Li+ is able to produce this effect. Preincubation of the soluble enzyme at low pH (5.5) and a change to a higher value (8.7-9.4) causes a decrease of the Hill coefficient with Li+ only; this effect is not detected using the homogenate. Our results suggest the following. (i) Li+ may neutralize negative charges of AChE more successfully than does Na+, resulting in higher activity, stabilization, and cooperativity of the enzyme. (ii) The KM values calculated at high substrate concentrations (greater than 200 microM) indicate that the substrate affinity of AChE can be increased only by Li+ binding on the enzyme. (iii) Changes in pH can modulate the cooperativity and may denature allosteric sites on the enzyme that bind Li+. (iv) Membrane, cations and (or) cellular factor(s) may regulate the cooperativity and substrate affinity of AChE, when they have been affected by pH changes.
Tsakiris
S
S
Department of Experimental Physiology, Medical School, University of Athens, Greece.
Kouniniotou-Krontiri
P
P
D000110:Acetylcholinesterase / Q000096:biosynthesis*; D000818:Animals; D003964:Diaphragm / Q000201:enzymology*; D004593:Electrophorus; D004790:Enzyme Induction; D004795:Enzyme Stability; D066298:In Vitro Techniques; D008094:Lithium / Q000494:pharmacology*; D011487:Protein Conformation; D051381:Rats; D012964:Sodium / Q000494:pharmacology*
D008094:Lithium; D012964:Sodium; D000110:Acetylcholinesterase