Substrate channeling of oxalacetate in solid-state complexes of malate dehydrogenase and citrate synthase.
Current evidence suggests that mitochondrial matrix enzymes exist in solid-state, multienzyme complexes in vivo. Addition of polyethylene glycol to a solution containing malate dehydrogenase and citrate synthase generates such a solid-state, enzyme complex in vitro at enzyme concentrations permitting kinetic measurements. Suspensions of the isolated, solid-state, hetero-complex of these enzymes were used to study the coupled reactions of citrate synthesis from malate, NAD, and CoASAc. The particles appear to be about 1 microgram in diameter. Considering the ratio of enzyme to oxalacetate molecules in or at the surface of the solid-state particles, one would expect oxalacetate to be converted to citrate within a few molecular distances of the site of oxalacetate generation. This model of "substrate channeling" (or alternatively a direct transfer of oxalacetate between enzymes) is supported by experiments with excess aspartate aminotransferase and glutamate added to the solution phase to give a reaction competing with the synthase for bulk phase oxalacetate. Quantities of aminotransferase that reduce the citrate reaction rate with soluble dehydrogenase and synthase by 90% do not significantly affect rates with comparable amounts of the dehydrogenase-synthase complex. We suggest that similar substrate channeling can occur in vivo and discuss the possible advantages provided thereby.
D000818:Animals; D002950:Citrate (si)-Synthase / Q000378:metabolism*; D002951:Citrates / Q000096:biosynthesis; D019343:Citric Acid; D003065:Coenzyme A / Q000378:metabolism; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000378:metabolism*; D008293:Malates / Q000378:metabolism; D008956:Models, Chemical; D009097:Multienzyme Complexes / Q000378:metabolism*; D009206:Myocardium / Q000201:enzymology; D009243:NAD / Q000378:metabolism; D010071:Oxaloacetates / Q000378:metabolism*; D007652:Oxo-Acid-Lyases / Q000378:metabolism*; D011092:Polyethylene Glycols; D013552:Swine
D002951:Citrates; D008293:Malates; D009097:Multienzyme Complexes; D010071:Oxaloacetates; D009243:NAD; D019343:Citric Acid; D011092:Polyethylene Glycols; C030298:malic acid; D008291:Malate Dehydrogenase; D002950:Citrate (si)-Synthase; D007652:Oxo-Acid-Lyases; D003065:Coenzyme A