Solute Effects on Biopolymer Processes

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DESCRIPTION (provided by applicant): In vitro, trial-and-error is used to select solutes to optimize or perturb biopolymer processes (e.g. folding, assembly, crystallization, binding). Our long term goal is to develop quantitative methods to predict or interpret solute effects on biopolymer processes in terms of the amounts and types of biopolymer surfaces buried or exposed. To this end, vapor pressure osmometry is being developed to measure interactions of denaturants, osmolytes, crystallization agents and Hofmeister salts with the relatively charged and polar surfaces of native proteins and nucleic acids. Interactions of these solutes with relatively uncharged, less polar biopolymer surfaces are quantified from their effects on selected biopolymer conformational changes (unfolding of marginally stable peptides and proteins; melting of nucleic acid helices). The quantitative information about interactions of solutes with types of charged, polar and nonpolar biopolymer surface obtained from this data will be used to predict or interpret solute effects in terms of structure. Results for urea and glycine betaine show the merit of this quantitative approach for two very different solutes. In vivo, amounts of cytoplasmic solutes and their interactions with biopolymers and solutes completely determine the amount of cytoplasmic water, volume, and osmolality. We observe large changes in the amount of cytoplasmic water and in cytoplasmic concentrations of biopolymers and solutes (e.g. [K+], [glycine betaine]) in E. coli upon shifts in osmolality or addition of osmoprotectants. For all conditions examined, these changes correlate with changes in growth rate. To test these correlations and to probe their molecular basis, cytoplasmic solute composition will be varied at constant osmolality using different osmoprotectants and using strains lacking one osmoprotectant porter. Effects on growth rate and on amounts of cytoplasmic water and solutes will be determined, and used in quantitative models of volume regulation and of compensation mechanisms to reduce the perturbing effects of changes in cytoplasmic [K+].