3-Aminoquinoline/p-Coumaric Acid as a MALDI Matrix for Glycopeptides, Carbohydrates, and Phosphopeptides

Abstract

Glycosylation and phosphorylation are important post-translational modifications in biological processes and biomarker research. The difficulty in analyzing these modifications is mainly their low abundance and dissociation of labile regions such as sialic acids or phosphate groups. One solution in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is to improve matrices for glycopeptides, carbohydrates, and phosphopeptides by increasing the sensitivity and suppressing dissociation of the labile regions. Recently, a liquid matrix 3-aminoquinoline (3-AQ)/α-cyano-4-hydroxycinnamic acid (CHCA) (3-AQ/CHCA), introduced by Kolli et al. in 1996, has been reported to increase sensitivity for carbohydrates or phosphopeptides, but it has not been systematically evaluated for glycopeptides. In addition, 3-AQ/CHCA enhances the dissociation of labile regions. In contrast, a liquid matrix 1,1,3,3-tetramethylguanidium (TMG, G) salt of p-coumaric acid (CA) (G3CA) was reported to suppress dissociation of sulfate groups or sialic acids of carbohydrates. Here we introduce a liquid matrix 3-AQ/CA for glycopeptides, carbohydrates, and phosphopeptides. All of the analytes were detected as [M + H](+) or [M - H](-) with higher or comparable sensitivity using 3-AQ/CA compared with 3-AQ/CHCA or 2,5-dihydroxybenzoic acid (2,5-DHB). The sensitivity was increased 1- to 1000-fold using 3-AQ/CA. The dissociation of labile regions such as sialic acids or phosphate groups and the fragmentation of neutral carbohydrates were suppressed more using 3-AQ/CA than using 3-AQ/CHCA or 2,5-DHB. 3-AQ/CA was thus determined to be an effective MALDI matrix for high sensitivity and the suppression of dissociation of labile regions in glycosylation and phosphorylation analyses.

Vibrational analysis of α-cyanohydroxycinnamic acid

“…This compound, which belongs to a group of hydroxycinnamic acid derivatives like sinapic/sinapinic, ferulic and caffeic acids, has been extensively used for protein and peptide analysis as a matrix for ultraviolet matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) [1,2]. Other commercially available cinnamic acids have been utilized in the analysis of proteins [1], oligodeoxyribonucleotides [3], glycopeptides [4], carbohydrates [4], and phosphopeptides [4]. Efforts to develop compounds with similar structures as sinapic acid and 4CHCA have been successful.…”

Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry: Mechanistic Studies and Methods for Improving the Structural Identification of Carbohydrates

“…As shown in Fig. 3, "cool" ionic liquid matrices: 1,1,3,3-tetramethylguanidium (TMG) salt of p-coumaric acid (G 3 CA), 107) and 3-aminoquinoline/pcoumaric acid (3-AQ/CA) 108,109) were developed by Tanaka and coworkers to yield a less locally heated ionization environment for enhancing the sensitivity and suppressing the losses of sialic acid moieties. Applying a similar concept, THAP-based ionic liquid matrices 110) were also reported for automated identi cation and quantitative analysis of native oligosaccharides.…”

Sensitive and Structure-Informative <i>N</i>-Glycosylation Analysis by MALDI-MS; Ionization, Fragmentation, and Derivatization

“…e liquid matrix 3-AQ/CHCA has now been largely replaced by the more versatile, sensitive, and "cold" liquid matrix 3-AQ/CA (3-aminoquinoline/p-coumaric acid). 25) An interesting feature of these on-target labeling methods is that they enable e ective negative-ion production from neutral glycans. In general, neutral glycans do not produce negative-ion species.…”