Optimizing protein crosslinking control: Synergistic quenching effects of glycine, histidine, and lysine on glutaraldehyde reactions

• Published in: Biochemical and biophysical research communications Vol. 702; p. 149567
• Main Authors: Kim, Kwang Sub, Lee, Yeseul, Lee, Ju Huck, Lee, Seung Sik, Chung, Jeong Min, Jung, Hyun Suk
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 02.04.2024
• Subjects: Cross-Linking Reagents - chemistry; Crosslinking reactions; Glutaral - chemistry; Glutaral - pharmacology; Glutaraldehyde; Glycine; Histidine; Lysine; Quenching activity1; Quenching efficiency; Quenching activity1; G; H; L; SEC; SDS-PAGE; Glutaraldehyde; Quenching efficiency; OD; LB; PRX2; GA; Crosslinking reactions; TEM; Quenching activity
• ISSN: 0006-291X; 1090-2104; 1090-2104
• DOI: 10.1016/j.bbrc.2024.149567

Glutaraldehyde (GA) is a protein crosslinker widely used in biochemical and pharmaceutical research because it can rapidly stabilize and immobilize substrates via amine group interactions. However, controlling GA crosslinking is challenging owing to its swift reactivity and the influence of various solution conditions, such as pH and concentrations of the substrate and crosslinker. Although extensive research has focused on GA cross-linking mechanisms, studies on quenching, which is critical for preventing non-specific aggregation during prolonged storage, remain sparse. This study examines the quenching efficiency of a combined amino acid mixture of glycine, histidine, and lysine, which are commonly used as individual quenchers. Our findings, confirmed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis, demonstrate that this amino acid blend offers superior quenching compared to single amino acids, enhancing quenching activity across a wide pH spectrum. These results provide a novel approach for mitigating the high reactivity of GA with implications for improving sample preservation and stabilization in a range of biochemical applications, including microscopy and cell fixation. •Quenching efficiency of a glycine, histidine, and lysine mixture was evaluated.•This combination offers superior quenching of glutaraldehyde fixation.•Quenching activity is enhanced across a wide pH spectrum.•The high reactivity of glutaraldehyde can thus be mitigated.