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Synthesizing Amino Acids Modified with Reactive Carbonyls in Silico to Assess Structural Effects Using Molecular Dynamics Simulations
Published on: April 26, 2024
In vitro study on structural alteration of myoglobin by methylglyoxal
Sauradipta Banerjee1, Abhay Sankar Chakraborti
1Department of Biophysics Molecular Biology and Bioinformatics, University of Calcutta, 92, Acharyya Prafulla Chandra Road, Kolkata, 700009, India.
Abstract:
Methylglyoxal (MG), a reactive α-oxoaldehyde, reacts with proteins to form irreversible advanced glycation end products (AGEs) following Maillard-like reaction. MG-induced AGE (MAGE) formation may be significant, particularly in diabetic condition with increased level of MG. Although myoglobin (Mb) is known to react with sugars to form AGEs, its interaction with MG is not known. Here we have studied interaction of Mb with MG. After in vitro reaction between Mb and MG at 25 °C for 7 days, the unchanged Mb and modified Mb (MG-Mb) were separated by ion exchange chromatography. Compared to Mb, MG-Mb exhibited higher electrophoretic mobility in native polyacrylamide gel electrophoresis, increased absorbance around 280 nm and more α-helical content, indicating structural changes of the modified protein. As shown by MALDI-mass spectrometry, MG converted Lys-16 and Lys-133 to carboxyethyllysine in MG-Mb. MAGE thus formed in MG-Mb may be associated with its enhanced mobility in native gel due to neutralization of positive charges and the observed structural changes in comparison with Mb.
Insights
Methylglyoxal (MG) reacts with myoglobin (Mb) to form modified myoglobin (MG-Mb), creating advanced glycation end products (AGEs). This modification alters protein structure and charge, with implications for conditions like diabetes.
Area of Science:
- Biochemistry
- Protein Chemistry
- Glycation Chemistry
Background:
- Methylglyoxal (MG) is a reactive α-oxoaldehyde implicated in forming irreversible advanced glycation end products (AGEs).
- MG-induced AGEs (MAGEs) are particularly relevant in diabetes due to elevated MG levels.
- While myoglobin (Mb) is known to form AGEs with sugars, its specific interaction with MG remains uncharacterized.
Purpose of the Study:
- To investigate the in vitro interaction between methylglyoxal (MG) and myoglobin (Mb).
- To characterize the structural and chemical modifications of myoglobin upon reaction with MG.
Main Methods:
- In vitro reaction of myoglobin (Mb) with methylglyoxal (MG) at 25°C for 7 days.
- Separation of modified myoglobin (MG-Mb) from native Mb using ion exchange chromatography.
- Analysis using native polyacrylamide gel electrophoresis (native PAGE), UV-Vis spectroscopy, and MALDI-mass spectrometry.
Main Results:
- MG-Mb exhibited increased electrophoretic mobility in native PAGE compared to native Mb.
- Modified myoglobin showed increased absorbance around 280 nm and enhanced α-helical content, indicating structural alterations.
- MALDI-mass spectrometry confirmed the conversion of lysine residues (Lys-16 and Lys-133) to carboxyethyllysine in MG-Mb.
Conclusions:
- Methylglyoxal (MG) modifies myoglobin (Mb) by forming MAGEs, specifically at Lys-16 and Lys-133.
- These modifications result in significant structural changes, including increased α-helical content and altered charge.
- The observed changes in MG-Mb, such as enhanced mobility, are attributed to charge neutralization and structural shifts.
