Charges in Hydrophobic Environments: A Strategy for Identifying Alternative States in Proteins

In the V23E variant of staphylococcal nuclease, Glu-23 has a pK a of 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 st...

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Bibliographic Details
Published in:Biochemistry (Easton) 2017-01, Vol.56 (1), p.212-218
Main Authors: Robinson, Aaron C, Majumdar, Ananya, Schlessman, Jamie L, García-Moreno E, Bertrand
Format: Article
Language:English
Subjects:
Crystallization
Crystallography, X-Ray
Glutamic Acid - chemistry
Glutamic Acid - genetics
Glutamic Acid - metabolism
Hydrogen-Ion Concentration
Hydrophobic and Hydrophilic Interactions
Kinetics
Magnetic Resonance Spectroscopy
Micrococcal Nuclease - chemistry
Micrococcal Nuclease - genetics
Micrococcal Nuclease - metabolism
Models, Molecular
Mutation, Missense
Protein Conformation
Protein Structure, Secondary
Staphylococcus aureus - enzymology
Staphylococcus aureus - genetics
Static Electricity
Thermodynamics
Valine - chemistry
Valine - genetics
Valine - metabolism
Water - chemistry
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Summary:In the V23E variant of staphylococcal nuclease, Glu-23 has a pK a of 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 pK a of an internal ionizable group can reflect the average of the very different pK a values (open ≈4.5, closed ≫7.5) sampled in the different conformational states. (2) The high apparent dielectric constant reported by the pK a value of internal ionizable group reflects conformational reorganization. (3) The apparent pK a of 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 pK a values of internal residues and to reproduce pH-driven conformational transitions of proteins.
ISSN:0006-2960
1520-4995
DOI:10.1021/acs.biochem.6b00843