Nature of the inactivation of the isocitrate dehydrogenase from an obligate halophile.
The nicotinamide adenine dinucleotide phosphate-specific isocitrate dehydrogenase (ICDH) of Halobacterium cutirubrum is rapidly inactivated at low NaCl levels. From sucrose gradient analysis, it was estimated that the active ICDH has an S(20,w) of 5.3 and a molecular weight of 75,000. The inactivation by removal of NaCl causes an unfolding of the protein yielding a less-compact conformer with an S(20,w) of 2.0. This inactivation apparently causes internal sulfhydryl groups to be exposed. Over 90% of the initial activity can be restored by dialyzing the inactivated ICDH against 4 m NaCl, provided that the exposed sulfhydryl groups are protected with dithiothreitol. The ICDH is permanently inactivated when the sulfhydryl groups are oxidized or alkylated. The alkylation of the inactive ICDH was demonstrated by treatment with (14)C-N-ethyl maleimide. Sucrose gradient analysis showed that (14)C was bound to a protein with sedimentation properties identical to that of reversibly inactivated ICDH, i.e., an S(20,w) of 2.0. Much less (14)C was bound when active ICDH was treated with (14)C-N-ethyl maleimide. The H. cutirubrum ICDH resembles other bacterial isocitrate dehydrogenases in being susceptible to concerted feedback inhibition by oxalacetate and glyoxalate.
D000577:Amides / Q000494:pharmacology; D000818:Animals; D002021:Buffers; D002247:Carbon Isotopes; D002499:Centrifugation, Density Gradient; D002712:Chlorides / Q000494:pharmacology; D002951:Citrates / Q000494:pharmacology; D003864:Depression, Chemical; D003956:Dialysis; D004789:Enzyme Activation; D005033:Ethylmaleimide / Q000494:pharmacology; D006038:Glyoxylates / Q000494:pharmacology; D006217:Halobacterium / Q000187:drug effects / Q000201:enzymology*; D007461:Iodoacetates / Q000494:pharmacology; D007521:Isocitrate Dehydrogenase / Q000378:metabolism*; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D009206:Myocardium / Q000201:enzymology; D009249:NADP; D010071:Oxaloacetates / Q000494:pharmacology; D012965:Sodium Chloride / Q000494:pharmacology; D013395:Sucrose; D013552:Swine
D000577:Amides; D002021:Buffers; D002247:Carbon Isotopes; D002712:Chlorides; D002951:Citrates; D006038:Glyoxylates; D007461:Iodoacetates; D010071:Oxaloacetates; D008345:Manganese; D012965:Sodium Chloride; D009249:NADP; D013395:Sucrose; D007521:Isocitrate Dehydrogenase; D005033:Ethylmaleimide