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Measuring protein-ligand interactions using liquid sample desorption electrospray ionization mass spectrometry.

ABSTRACT: We have previously shown that liquid sample desorption electrospray ionization-mass spectrometry (DESI-MS) is able to measure large proteins and noncovalently bound protein complexes (to 150 kDa) (Ferguson et al., Anal. Chem. 2011, 83, 6468-6473). In this study, we further investigate the application of liquid sample DESI-MS to probe protein-ligand interactions. Liquid sample DESI allows the direct formation of intact protein-ligand complex ions by spraying ligands toward separate protein sample solutions. This type of "reactive" DESI methodology can provide rapid information on binding stiochiometry, selectivity, and kinetics, as demonstrated by the binding of ribonuclease A (RNaseA, 13.7 kDa) with cytidine nucleotide ligands and the binding of lysozyme (14.3 kDa) with acetyl chitose ligands. A higher throughput method for ligand screening by liquid sample DESI was demonstrated, in which different ligands were sequentially injected as a segmented flow for DESI ionization. Furthermore, supercharging to enhance analyte charge can be integrated with liquid sample DESI-MS, without interfering with the formation of protein-ligand complexes.

SUBMITTER: Liu P

PROVIDER: S-EPMC3901310 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

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Measuring protein-ligand interactions using liquid sample desorption electrospray ionization mass spectrometry.

Liu Pengyuan P Zhang Jiang J Ferguson Carly N CN Chen Hao H Loo Joseph A JA

Analytical chemistry 20131122 24

We have previously shown that liquid sample desorption electrospray ionization-mass spectrometry (DESI-MS) is able to measure large proteins and noncovalently bound protein complexes (to 150 kDa) (Ferguson et al., Anal. Chem. 2011, 83, 6468-6473). In this study, we further investigate the application of liquid sample DESI-MS to probe protein-ligand interactions. Liquid sample DESI allows the direct formation of intact protein-ligand complex ions by spraying ligands toward separate protein sample ...[more]

PMID: 24237005

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Project description:We recently reported the use of desorption electrospray ionization (DESI) as a novel interface to couple high-performance liquid chromatography (HPLC) with mass spectrometry (MS) (Chem. Commun. 2011, 47, 4171). One of the benefits of such an interface is that post-column derivatization of separated analytes can be integrated with ionization via a "reactive" DESI approach in which a derivatizing reagent is doped into the spray solvent. The reactive DESI interface allows analyte desorption/ionization from the end of the chromatographic column with prompt MS detection; a short time delay of ~20 ms was demonstrated. In this study, we extended this application by "supercharging" proteins following HPLC separation using a DESI spray solvent containing supercharging reagents, m-nitrobenzyl alcohol (m-NBA) or sulfolane. Proteins (insulin, ubiquitin, lysozyme and ?-lactalbumin) eluted out of the LC column can be supercharged with the protein charge state distributions (CSDs) significantly increased (to higher charge), which would be advantageous for subsequent top-down MS analysis of proteins. Interestingly, supercharging combined with reactive DESI enhances tolerance towards trifluoroacetic acid (TFA), which is known to be a superior additive in the mobile phase for premium peptide/protein chromatographic separation but has severe signal suppression effects for conventional electrospray ionization (ESI). In comparison to electrosonic spray ionization (ESSI), a variant form of ESI, the sensitivity of protein analysis using LC/DESI-MS with the mobile phase containing TFA can be improved by up to 70-fold for lysozyme and ?-lactalbumin by including m-NBA in the DESI spray solvent. Presumably, by reducing TFA dissociation in the droplet, supercharging agents lower trifluoroacetate anion concentrations and concomitantly reduce ion pairing to analyte cationic sites. The reduced ion pairing therefore decreases the TFA signal suppression effect. The supercharging capability and the reduction of TFA signal suppression suggest that LC/DESI-MS is a valuable method for protein analysis.

Dataset Information

Measuring protein-ligand interactions using liquid sample desorption electrospray ionization mass spectrometry.

Publications

Measuring protein-ligand interactions using liquid sample desorption electrospray ionization mass spectrometry.

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