Organization-dependent effects of toxic bivalent ions microtubule assembly and glycolysis.
The effects of bivalent ions on tubulin dynamics and the upper phase of glycolysis were investigated at different organization levels in vitro. Cu2+, Cd2+, Hg2+ and CrO4(2-) inhibit the tubulin polymerization at an IC50 of 14-24 microM with high cooperativity and also induce microtubule disassembly. The apparent binding constants of the ions to tubulin, estimated by fluorescence quenching, vary between 6 and 28 microM. BIAcore measurements for tubulin-tubulin interaction suggest that the presence of Cu2+ affects neither koff nor kon, but the amount of the bound tubulin. While the inhibitory effect of Cu2+ on tubulin polymerization is partially abolished by cross-linking of microtubules with substoichiometric amounts of phosphofructokinase or decoration of tubules with cytosolic proteins, in the presence of kinase but not with cytosolic proteins the tubules are resistant to CrO4(2-). No inhibitory effect of Cu2+ or CrO4(2-) on microtubule assembly was detected in the MAP-containing cytosolic fraction. Electron microscopy revealed that tubules assembled in the presence of Cu2+ or CrO4(2-) ions contain aggregates of thread-like oligomers that are less conspicuous in the presence of cytosolic proteins. Cu2+, Cd2+, and Hg2+ inhibit the glycolytic flux in the cytosolic fraction characterized at equilibrium by an IC50 of 10-14 microM with high cooperativity. Tubulin diminishes the inhibitory effect of the cations. These data indicate that the responses elicited by the bivalent ions are highly dependent on the supramolecular organization of the systems.
Liliom
K
K
Institute of Enzymology, Biological Research Center of the Hungarian Academy of Sciences, Budapest, Hungary.
Wágner
G
G
Pácz
A
A
Cascante
M
M
Kovács
J
J
Ovádi
J
J
D000085:Acetates / Q000494:pharmacology; D000818:Animals; D001921:Brain / Q000378:metabolism; D002104:Cadmium / Q000494:pharmacology; D002417:Cattle; D002840:Chromates / Q000494:pharmacology; D003300:Copper / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D006019:Glycolysis; D020128:Inhibitory Concentration 50; D007477:Ions; D007700:Kinetics; D017669:Mercury Compounds / Q000494:pharmacology; D008854:Microscopy, Electron; D008870:Microtubules / Q000378:metabolism* / Q000502:physiology* / Q000648:ultrastructure; D009566:Nitrates / Q000494:pharmacology; D009942:Organometallic Compounds / Q000494:pharmacology; D010732:Phosphofructokinase-1 / Q000378:metabolism; D017680:Potassium Compounds / Q000494:pharmacology; D011485:Protein Binding; D012977:Sodium Nitrite / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D020349:Surface Plasmon Resonance; D013997:Time Factors
D000085:Acetates; D002840:Chromates; D007477:Ions; D017669:Mercury Compounds; D009566:Nitrates; D009942:Organometallic Compounds; D017680:Potassium Compounds; D002104:Cadmium; C029948:mercury nitrate; C027373:potassium chromate(VI); D003300:Copper; C028031:cadmium acetate; D010732:Phosphofructokinase-1; D012977:Sodium Nitrite; C008261:lead acetate; C029892:cupric chloride