MOLYBDOPTERIN GUANINE DINUCLEOTIDE IS THE ORGANIC MOIETY OF THE MOLYBDENUM COFACTOR IN RESPIRATORY NITRATE REDUCTASE FROM PSEUDOMONAS-STUTZERI

Respiratory nitrate reductase from the denitrifying bacterium Pseudomo nas stutzeri is an iron-sulfur enzyme containing the molybdenum cofact or. Hydrolysis of native nitrate reductase with aqueous sulfuric acid revealed 0.92 mol of 5'-GMP per mol of enzyme. The pterin present in t he molybdenum cofactor was liberated from the protein and reacted with iodoacetamide. The resulting di(carboxamidomethyl) (cam) derivative w as purified on a C18-cartridge and analyzed for its structural element s. Treatment of the cam derivative with nucleotide pyrophosphatase and subsequent HPLC analysis revealed the formation of di(cam)molybdopter in and 5'-GMP at a 1:1 molar ratio and with a yield of 79% with respec t to the molybdenum content of the enzyme. Treatment of the cam deriva tive with nucleotide pyrophosphatase and alkaline phosphatase led to t he liberation of 0.51 mol dephosphodi(cam)molybdopterin and of 0.59 mo l guanosine per mol of enzyme, which is equal to a molar ratio of 1:1. 2. The results indicate, that the organic moiety of the molybdenum cof actor of nitrate reductase from P. stutzeri is molybdopterin guanine d inucleotide of which one mol is contained per mol of nitrate reductase .