The relationship of Mo, molybdopterin, and the cyanolyzable sulfur in the Mo cofactor.
Reconstitution of the apoprotein of the molybdoenzyme nitrate reductase in extracts of the Neurospora crassa mutant nit-1 with molybdenum cofactor released by denaturation of purified molybdoenzymes is efficient in the absence of exogenous MoO2-4 under defined conditions. Evidence is presented that this molybdate-independent reconstitution is due to transfer of intact Mo cofactor, a complex of Mo and molybdopterin (MPT), the organic constituent of the cofactor. This complex can be separated from denatured protein by gel filtration, and from excess MoO2-4 by reverse-phase HPLC. Sulfite oxidase, native xanthine dehydrogenase, and cyanolyzed xanthine dehydrogenase are equipotent Mo cofactor donors. Other well-studied inactive forms of xanthine dehydrogenase are also shown to be good cofactor sources. Using xanthine dehydrogenase specifically radiolabeled in the cyanolyzable sulfur, it is shown that this terminal ligand of Mo is rapidly removed from Mo cofactor under the conditions used for reconstitution.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D003067:Coenzymes; D005033:Ethylmaleimide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000201:enzymology; D008628:Mercury / Q000494:pharmacology; D008667:Metalloproteins; D008982:Molybdenum / Q000378:metabolism*; D000090006:Molybdenum Cofactors; D009492:Neurospora crassa / Q000235:genetics; D050895:Nitrate Reductase (NADH); D009565:Nitrate Reductases / Q000378:metabolism; D011489:Protein Denaturation; D011621:Pteridines / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000032:analysis; D013440:Sulfides / Q000032:analysis; D014414:Tungsten / Q000494:pharmacology; D017973:Tungsten Compounds; D014968:Xanthine Dehydrogenase / Q000378:metabolism
D003067:Coenzymes; D005033:Ethylmaleimide; D008628:Mercury; D008667:Metalloproteins; D008982:Molybdenum; D000090006:Molybdenum Cofactors; D050895:Nitrate Reductase (NADH); D009565:Nitrate Reductases; D011621:Pteridines; D013438:Sulfhydryl Compounds; D013440:Sulfides; D014414:Tungsten; D017973:Tungsten Compounds; D014968:Xanthine Dehydrogenase; C022261:molybdopterin; C021101:tungstic(VI) acid