Related Experiment Video
Updated: Aug 26, 2026
Analysis of β-Amyloid-induced Abnormalities on Fibrin Clot Structure by Spectroscopy and Scanning Electron Microscopy
Published on: November 30, 2018
Laser light-scattering evidence for an altered association of beta B1-crystallin deamidated in the connecting peptide
Michael J Harms1, Philip A Wilmarth, Deborah M Kapfer
1Oregon Health and Science University, Portland, OR 97239, USA.
Abstract:
Deamidation is a prevalent modification of crystallin proteins in the vertebrate lens. The effect of specific sites of deamidation on crystallin stability in vivo is not known. Using mass spectrometry, a previously unreported deamidation in beta B1-crystallin was identified at Gln146. Another deamidation was investigated at Asn157. It was determined that whole soluble beta B1 contained 13%-17% deamidation at Gln146 and Asn157. Static and quasi-elastic laser light scattering, circular dichroism, and heat aggregation studies were used to explore the structure and associative properties of recombinantly expressed wild-type (wt) beta B1 and the deamidated beta B1 mutants, Q146E and N157D. Dimer formation occurred for wt beta B1, Q146E, and N157D in a concentration-dependent manner, but only Q146E showed formation of higher ordered oligomers at the concentrations studied. Deamidation at Gln146, but not Asn157, led to an increased tendency of beta B1 to aggregate upon heating. We conclude that deamidation creates unique effects depending upon where the deamidation is introduced in the crystallin structure.
Insights
Deamidation of beta B1-crystallin at Gln146, but not Asn157, increases protein aggregation. This highlights how specific deamidation sites impact crystallin stability and lens protein structure.
Area of Science:
- Biochemistry
- Structural Biology
- Ophthalmology
Background:
- Deamidation is a common post-translational modification in lens crystallins.
- The impact of deamidation site-specificity on crystallin stability in vivo remains unclear.
Purpose of the Study:
- To investigate the structural and stability effects of specific deamidation sites in beta B1-crystallin.
- To determine if deamidation at Gln146 and Asn157 influences beta B1-crystallin aggregation and oligomerization.
Main Methods:
- Mass spectrometry to identify deamidation sites.
- Recombinant expression of wild-type and deamidated beta B1-crystallin mutants (Q146E, N157D).
- Static/quasi-elastic laser light scattering, circular dichroism, and heat aggregation assays to assess protein structure and properties.
Main Results:
- Deamidation at Gln146 (Q146E) induced higher-order oligomer formation, unlike wild-type or N157D mutants.
- Deamidation at Gln146 significantly increased beta B1-crystallin aggregation upon heating.
- Deamidation at Asn157 did not show similar effects on aggregation or oligomerization.
Conclusions:
- Deamidation's effect on beta B1-crystallin stability is site-dependent.
- Gln146 deamidation promotes aggregation, suggesting a role in age-related lens opacities.
- Understanding site-specific modifications is crucial for comprehending protein behavior in biological systems.
More Related Videos
09:31Visualization of Amyloid β Deposits in the Human Brain with Matrix-assisted Laser Desorption/Ionization Imaging Mass Spectrometry
Published on: March 7, 2019
10:36Covalent Labeling with Diethylpyrocarbonate for Studying Protein Higher-Order Structure by Mass Spectrometry
Published on: June 15, 2021