Sequence Reversal Prevents Chain Collapse and Yields Heat-Sensitive Intrinsic Disorder.

Sequence patterns of charge, hydrophobicity, hydrogen bonding, and other amino acid physicochemical properties contribute to mechanisms of protein folding, but how sequence composition and patterns influence the conformational dynamics of the denatured state ensemble is not fully understood. To investigate structure-sequence relationships in the denatured state, we reversed the sequence of staphylococcal nuclease and characterized its structure, thermodynamic character, and hydrodynamic radius using circular dichroism spectroscopy, dynamic light scattering, analytical ultracentrifugation, and size-exclusion chromatography as a function of temperature. The macromolecular size of "Retro-nuclease" is highly expanded in solution with characteristics similar to biological intrinsically disordered proteins.

Next-Generation AUC: Analysis of Multiwavelength Analytical Ultracentrifugation Data.

We describe important advances in methodologies for the analysis of multiwavelength data. In contrast to the Beckman-Coulter XL-A/I ultraviolet-visible light detector, multiwavelength detection is able to simultaneously collect sedimentation data for a large wavelength range in a single experiment. The additional dimension increases the data density by orders of magnitude, posing new challenges for data analysis and management.

A parametrically constrained optimization method for fitting sedimentation velocity experiments.

A method for fitting sedimentation velocity experiments using whole boundary Lamm equation solutions is presented. The method, termed parametrically constrained spectrum analysis (PCSA), provides an optimized approach for simultaneously modeling heterogeneity in size and anisotropy of macromolecular mixtures. The solutions produced by PCSA are particularly useful for modeling polymerizing systems, where a single-valued relationship exists between the molar mass of the growing polymer chain and its corresponding anisotropy.