MULTIPLE CONFORMATIONAL STATES OF A NEW HEMATOPOIETIC CYTOKINE (MEGAKARYOCYTE GROWTH AND DEVELOPMENT FACTOR) - PH-INDUCED AND UREA-INDUCED DENATURATION

The effect of pH and urea on the conformation of recombinant human meg akaryocyte growth and development factor (rHuMGDF) was determined by c ircular dichroism, intrinsic fluorescence spectroscopy, and equilibriu m ultracentrifugation, The conformation of rHuMGDF was dependent on pH and urea concentration. Multiple folding forms were evidenced by mult iple pH-induced transitions and urea-induced equilibrium transitions t hat deviated from a simple two-state process. In neutral to alkaline p H, rHuMGDF exists as a monomer, but an acid-induced conformational sta te self-associates to form a soluble aggregate. A folding intermediate (s) was observed with a more stable secondary structure than tertiary structure and was dependent on the pH of the urea-induced denaturation , The differences in the stabilities of the folding states were most d istinct in the pH range of 4.5 to 6.5, The presence of intermediates i n the folding pathway of rHuMGDF are similar to findings of previous s tudies of related growth factors that share a common three-dimensional structure. (C) 1998 Wiley-Liss, Inc.