PREFERENTIAL EXCLUSION OF SUCROSE FROM RECOMBINANT INTERLEUKIN-1 RECEPTOR ANTAGONIST - ROLE IN RESTRICTED CONFORMATIONAL MOBILITY AND COMPACTION OF NATIVE-STATE

Understanding the mechanism for sucrose-induced protein stabilization is important in many diverse fields, ranging from biochemistry and env ironmental physiology to pharmaceutical science. Timasheff and Lee [Le e, J. C. & Timasheff, S. N. (1981) J. Biol. Chem, 256, 7193-7201] have established that thermodynamic stabilization of proteins by sucrose i s due to preferential exclusion of the sugar from the protein's surfac e, which increases protein chemical potential, The current study measu res the preferential exclusion of 1 M sucrose from a protein drug, rec ombinant interleukin 1 receptor antagonist (rhIL-1ra), It is proposed that the degree of preferential exclusion and increase in chemical pot ential are directly proportional to the protein surface area and that, hence, the system will favor the protein state with the smallest surf ace area, This mechanism explains the observed sucrose-induced restric tion of rhIL-1ra conformational fluctuations, which were studied by hy drogen-deuterium exchange and cysteine reactivity measurements, Furthe rmore, infrared spectroscopy of rhlL-1ra suggested that a more ordered native conformation is induced by sucrose, Electron paramagnetic reso nance spectroscopy demonstrated that in the presence of sucrose, spin- labeled cysteine 116 becomes more buried in the protein's interior and that the hydrodynamic diameter of the protein is reduced, The prefere ntial exclusion of sucrose from the protein and the resulting shift in the equilibrium between protein states toward the most compact confor mation account for sucrose-induced effects on rhIL-1ra.