Direct Measurement and Computation of Forces Between Membranes or Macromolecules

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r thoughts this year have ranged from the intimacies of specific protein/DNA contact to the grand symmetries of DNA packed into macroscopic liquid-crystal assembly. There are qualitative differences in the amount of water trapped between protein and DNA either loosely and non-specifically associated or tightly and dryly packed; a difference of 70 water molecules can occur because of a single mutation among six base pairs. DNA packed at the density seen in viral capsids shows a hitherto unobserved "hexatic" order wherein the angular arrangement is that of a hexagonal lattice two-dimensional crystal while the distances between molecules have liquid-like rather than crystalline order. Our Osmotic Stress procedure for condensing DNA to high concentrations where it is examined by x-ray diffraction is a procedure for measuring forces as well as the work or 'free energy' needed to create structures. We have developed and successfully tested a model of DNA-DNA interaction where the molecules collide through long-range repulsive forces without physical contact. Similar measurements on D- and L-form polypeptides have led us to think about the connection between molecular 'chirality' or twist and molecular packing. Because drug specificity is sometimes based on chirality, there are practical consequences to this feature of molecular structure. Measurements on polysaccharide interactions show the same kind of exponentially varying 'hydration forces' that we have discovered to dominate the interaction of all large biomolecules approaching contact. Further evidence that these forces are due to restructuring of water around the surface comes from our measurement of NaCl exclusion by polysaccharides; solubility of salt drops off with an exponential dependence of the same characteristic length seen in the forces. Besides this, heating the polysaccharide/water phases causes water to be released, as though ordered water were being melted off the chains. Similar measurements on sickle-cell hemoglobin gels also show gel shrinkage with heating; in this case we have been able to measure the positive change in entropy, or molecular disorder, that goes with condensation of the gel. The same osmotic-stressing logic used by us for force measurement is applied to the problem of protein crystallization. We have been able to measure the entropy and enthalpy of lysozyme crystallization; we expect to use this information to modify crystallization conditions systematically to improve methods of preparation for structure determination. Theoretical work has concentrated on the statistical mechanics of ions in the presence of polyelectrolytes such as DNA, charged bilayers, or large colloidal particles. We have been able to re-formulate ionic-fluctuation forces to take account of boundary surfaces where forces have been seen to differ from those seen in bulk solution. We have developed a model for averaging salt concentration over the millisecond-lifetimes of proteins' functional states in order to derive quantitative connections between salt activity, solution conductance near charged surfaces, and the free energy of charged molecules. Linear charged molecules will change their flexibility in the presence of high salt; they will also show buckling instabilities where the molecules can collapse on themselves." " We have developed a systematic formalism that is allowing us to design experiments to examine these changes with salt condition. Our association with NASA has now passed its second year where we tested different x-ray lenses with materials prepared in our lab. The results suggest that accurate force measurement can be carried out with the NASA lens that can boost x-ray flux by factors of 10 to 100 times."