Reduction of Cys(36)-Cys(42) and Cys(64)-Cys(74) disulfide bonds in recombinant human granulocyte colony stimulating factor

The Cys(36)-Cys(42) and Cys(64)-Cys(74) disulfide bonds in recombinant meth ionyl human granulocyte colony-stimulating factor were reduced to sulfhydry ls with dithiothreitol or mercury. Both reduction reactions are dependent o n the pH. The reduction reaction with dithiothreitol increased in rate with increasing pH; between pH 7-9 and above pH 10.5 this increase was less tha n in other regions. These observations are explained by repulsive forces be tween dithiothreitol and regions in granulocyte colony-stimulating factor w hich intensify in these pH-regions. The hydroxyl catalysis causes the overa ll increase in k(obs) in the pH-region studied. The reduction of the disulf ides with mercury is, as could be expected from the Nernst equation for dis ulfide reduction, also pH dependent: the half-wave potential decreases with increasing pH as predicted by theory. (C) 1999 Elsevier Science B.V. All r ights reserved.