TitleDifferences in solution dynamics between lens β-crystallin homodimers and heterodimers probed by hydrogen-deuterium exchange and deamidation.
Publication TypeJournal Article
Year of Publication2016
AuthorsLampi KJ, Murray MR, Peterson MP, Eng BS, Yue E, Clark AR, Barbar E, David LL
JournalBiochim Biophys Acta
Issue1 Pt B
Date Published2016 Jan
KeywordsAmides, Amino Acid Sequence, Animals, beta-Crystallins, Binding Sites, Deuterium Exchange Measurement, Dimerization, Humans, Lens, Crystalline, Molecular Probe Techniques, Molecular Sequence Data, Protein Binding, Protein Conformation, Protein Multimerization

BACKGROUND: Lens transparency is due to the ordered arrangement of the major structural proteins, called crystallins. βB2 crystallin in the lens of the eye readily forms dimers with other β-crystallin subunits, but the resulting heterodimer structures are not known and were investigated in this study.

METHODS: Structures of βA3 and βB2 crystallin homodimers and the βA3/βB2 crystallin heterodimers were probed by measuring changes in solvent accessibility using hydrogen-deuterium exchange with mass spectrometry. We further mimicked deamidation in βB2 and probed the effect on the βA3/βB2 heterodimer. Results were confirmed with chemical crosslinking and NMR.

RESULTS: Both βA3 and βB2 had significantly decreased deuterium levels in the heterodimer compared to their respective homodimers, suggesting that they had less solvent accessibility and were more compact in the heterodimer. The compact structure of βB2 was supported by the identification of chemical crosslinks between lysines in βB2 within the heterodimer that were inconsistent with βB2's extended homodimeric structure. The compact structure of βA3 was supported by an overall decrease in mobility of βA3 in the heterodimer detected by NMR. In βB2, peptides 70-84 and 121-134 were exposed in the homodimer, but buried in the heterodimer with ≥50% decreases in deuterium levels. Homologous peptides in βA3, 97-109 and 134-149, had 25-50% decreases in deuterium levels in the heterodimer. These peptides are probable sites of interaction between βB2 and βA3 and are located at the predicted interface between subunits with bent linkers. Deamidation at Q184 in βB2 at this predicted interface led to a less compact βB2 in the heterodimer. The more compact structure of the βA3/βB2 heterodimer was also more heat stable than either of the homodimers.

CONCLUSIONS: The major structural proteins in the lens, the β-crystallins, are not static, but dynamic in solution, with differences in accessibility between the homo-and hetero-dimers. This structural flexibility, particularly of βB2, may facilitate formation of different size higher-ordered structures found in the transparent lens.

GENERAL SIGNIFICANCE: Understanding complex hetero-oligomer interactions between β-crystallins in normal lens and how these interactions change during aging is fundamental to understanding the cause of cataracts. This article is part of a Special Issue entitled Crystallin Biochemistry in Health and Disease.

Alternate JournalBiochim. Biophys. Acta
PubMed ID26145577
PubMed Central IDPMC4673030
Grant ListR01 EY007755 / EY / NEI NIH HHS / United States
GM084276-01 / GM / NIGMS NIH HHS / United States
P30 EY010572 / EY / NEI NIH HHS / United States
EY007755 / EY / NEI NIH HHS / United States
R29 EY012239 / EY / NEI NIH HHS / United States
R01 GM084276 / GM / NIGMS NIH HHS / United States
EY012239 / EY / NEI NIH HHS / United States
EY10572 / EY / NEI NIH HHS / United States
R01 EY012239 / EY / NEI NIH HHS / United States