title: Charges in Hydrophobic Environments: A Strategy for Identifying Alternative States in Proteins.
abstract: In the V23E variant of staphylococcal nuclease, Glu-23 has a pKaof 7.5. At low pH, Glu-23 is neutral and buried in the hydrophobic interior of the protein. Crystal structures and NMR spectroscopy experiments show that when Glu-23 becomes charged, the protein switches into an open state in which strands β1 and β2 separate from the β-barrel; the remaining structure is unaffected. In the open state the hydrophobic interior of the protein is exposed to bulk water, allowing Glu-23 to become hydrated. This illustrates several key aspects of protein electrostatics: (1) The apparent pKaof an internal ionizable group can reflect the average of the very different pKavalues (open ≈4.5, closed ≫7.5) sampled in the different conformational states. (2) The high apparent dielectric constant reported by the pKavalue of internal ionizable group reflects conformational reorganization. (3) The apparent pKaof internal groups can be governed by large conformational changes. (4) A single charge buried in the hydrophobic interior of a protein is sufficient to convert what might have been a transient, partially unfolded state into the dominant state in solution. This suggests a general strategy for examining inaccessible regions of the folding landscape and for engineering conformational switches driven by small changes in pH. These data also constitute a benchmark for stringent testing of the ability of computational algorithms to predict pKavalues of internal residues and to reproduce pH-driven conformational transitions of proteins.
authors.forename: Aaron C
authors.identifier: http://orcid.org/0000-0001-6410-7147
authors.initials: AC
authors.lastname: Robinson
authors.forename: Ananya
authors.initials: A
authors.lastname: Majumdar
authors.affiliation: Chemistry Department, United States Naval Academy , 572M Holloway Rd MS 9B, Annapolis, Maryland 21402, United States.
authors.forename: Jamie L
authors.initials: JL
authors.lastname: Schlessman
authors.forename: Bertrand
authors.initials: B
authors.lastname: García-Moreno E
mesh_terms: D003460:Crystallization; D018360:Crystallography, X-Ray; D018698:Glutamic Acid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008836:Micrococcal Nuclease / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D020125:Mutation, Missense; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013211:Staphylococcus aureus / Q000201:enzymology / Q000235:genetics; D055672:Static Electricity; D013816:Thermodynamics; D014633:Valine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014867:Water / Q000737:chemistry
chemical_list: D014867:Water; D018698:Glutamic Acid; D008836:Micrococcal Nuclease; D014633:Valine