Comparison of Protein Structure in the Crystal and in Solution. I. The Tyrosyl Ionization of Crystalline Methemoglobin^*

Horse Heart Cytochrome c. Spectrophotometric Titration and Viscosity Changes in Alkaline Solution*

“…The High-pH Difference Spectrum of Cytochrome c.-The ultraviolet difference spectrum of cytochrome c at pH 11.5 against a pH 8.9 reference is shown in Figure 1, and is characteristic of the tyrosyl ionization (Hermans, 1962; Wetlaufer, 1962). The maximum at 242 µ is shifted to the blue of the corresponding 245µ maximum in hemoglobin (Hermans, 1962;Rupley, 1964). The intensity ratio of the 242to the 295-mp peak is 4.32, similar to the values found for tyrosine (4.25) and methemoglobin (4.47) (Hermans, 1962;Rupley, 1964).…”

“…The maximum at 242 µ is shifted to the blue of the corresponding 245µ maximum in hemoglobin (Hermans, 1962;Rupley, 1964). The intensity ratio of the 242to the 295-mp peak is 4.32, similar to the values found for tyrosine (4.25) and methemoglobin (4.47) (Hermans, 1962;Rupley, 1964). Spectrophotometric Titration Curve.-Figure 2 shows the change in ionization of the four tyrosine residues of horse heart cytochrome c as a function of pH.…”

Recent advances and applications of diffuse X-ray small-angle scattering on biopolymers in dilute solutions

“…It should be mentioned that Rupley (1968) carried out interesting investigations by other methods in order to determine how closely the structure in solution resembles that in the crystal. With this aim in view he studied, above all, changes in protein solubility (Rupley & Gates, 1968), changes in proton binding and in volume (Krivacic & Rupley, 1968) at the transition of the protein into the crystalline state. As a result of these investigations he assumes that in haemoglobin, for instance, there can be no essential conformation difference between the molecule in crystal and in solution.…”

Characterizing Enzyme Reactions in Microcrystals for Effective Mix-and-Inject Experiments using X-ray Free-Electron Lasers

“…However, there are currently limited options to assess these quantities in crystallo. Solution assays provide some guidance for optimizing conditions, but reaction rates in crystals and solutions can vary by several orders of magnitude. − These differences may result from ligand diffusion times, interference from crystal contacts, limited active site accessibility, or buffer chemistry. Kinetics can also vary for different crystallization conditions for the same biomolecule.…”

“…While previous experiments have characterized reactions in microcrystals using a stopped-flow mixer, − the mixing methods and delay mechanisms associated with this technique differ significantly from those of current XFEL mixing injectors. Therefore, timescales may not transfer between these devices.…”