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Strong hydrogen bonding interactions involving a buried glutamic acid in the transmembrane sequence of the neu/erbB-2
S O Smith1, C S Smith, B J Bormann
1Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520-8114, USA.
Abstract:
The receptor tyrosine kinase encoded by the neu/erbB-2 proto-oncogene is constitutively activated by a single valine to glutamic acid substitution at position 664 in the predicted membrane-spanning sequence of the receptor. We have explored the structural changes involved in receptor activation with polarized FTIR and magic angle spinning NMR spectroscopy. The hydrophobic transmembrane sequence folds into a well-defined alpha-helical structure spanning the membrane bilayer. Measurements of the pKa and 13C chemical shift anisotropy of Glu 664 reveal that the side chain carboxyl group is protonated and strongly hydrogen bonded. These studies provide direct evidence for glutamate hydrogen-bonding interactions in the mechanism of receptor dimerization and activation.
Insights
The neu/erbB-2 receptor tyrosine kinase is activated by a mutation causing glutamate to form hydrogen bonds. This structural change in the transmembrane helix drives receptor dimerization and activation.
Area of Science:
- Biochemistry
- Molecular Biology
- Structural Biology
Background:
- The neu/erbB-2 proto-oncogene encodes a receptor tyrosine kinase.
- Constitutive activation of this receptor is linked to specific mutations within its transmembrane domain.
Purpose of the Study:
- To investigate the structural alterations underlying neu/erbB-2 receptor activation.
- To elucidate the role of the transmembrane sequence and specific mutations in receptor function.
Main Methods:
- Utilized polarized Fourier Transform Infrared (FTIR) spectroscopy.
- Employed magic angle spinning Nuclear Magnetic Resonance (NMR) spectroscopy.
- Measured the pKa and 13C chemical shift anisotropy of Glutamate 664.
Main Results:
- The transmembrane sequence adopts a stable alpha-helical structure.
- Glutamate 664's side chain carboxyl group is protonated and forms strong hydrogen bonds.
- These findings indicate specific glutamate interactions are crucial for activation.
Conclusions:
- The alpha-helical transmembrane domain plays a key role in receptor activation.
- Glutamate hydrogen-bonding interactions are directly implicated in receptor dimerization and signaling.
- Structural insights into oncogenic receptor activation mechanisms were provided.