title: Effects of a naturally occurring compatible osmolyte on the internal dynamics of ribonuclease A.
abstract: Osmolytes are small organic solutes accumulated intracellularly by many organisms as they adapt to environmental stresses. Compatible osmolytes, a functional class of osmolytes, increase protein stability while having little or no effect on protein function. To investigate the interrelationships between protein stability, function, and internal dynamics, a hydrogen exchange (HX) quench method was established and used to study the effects of sucrose (a typical compatible osmolyte) on the structural fluctuations of ribonuclease A. It was found that the HX rates of the amide protons with intermediate rates are not affected by 1 M sucrose, but the slow-exchanging amide protons exchange even slower in 1 M sucrose. The protection factors of the slow-exchanging protons fall into a comparatively narrow range while those of the intermediate-exchanging protons vary widely. In agreement with the two-process model [Woodward, C.K., & Hilton, B. D. (1980) Biophys. J. 32, 561-575], we conclude that for those slow-exchanging amide protons, the exchange occurs mainly from the compact unfolded state ensemble of the protein. The internal dynamics leading to slow exchange involve exposure of large protein surface areas, similar to that which occurs upon the unfolding of protein. Because sucrose opposes such an increase in protein surface area exposure, both the slow HX rates and the protein stability are affected by sucrose. For those amide protons with fast and intermediate HX rates, the exchange occurs mainly from the native state ensemble of the protein. The internal dynamics involved in the exchange are localized without much surface area change, and functionally important structural fluctuations are likely to occur within this dynamic range.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Biochemistry, Southern Illinois University at Carbondale 62901, USA.
authors.forename: A
authors.initials: A
authors.lastname: Wang
authors.forename: A D
authors.initials: AD
authors.lastname: Robertson
authors.forename: D W
authors.initials: DW
authors.lastname: Bolen
mesh_terms: D000818:Animals; D002417:Cattle; D003903:Deuterium; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D010179:Pancreas / Q000201:enzymology; D011522:Protons; D012259:Ribonuclease, Pancreatic / Q000737:chemistry* / Q000187:drug effects; D013395:Sucrose / Q000494:pharmacology*
chemical_list: D011522:Protons; D013395:Sucrose; D003903:Deuterium; D012259:Ribonuclease, Pancreatic