Screening for Noncovalent Ligand−Receptor Interactions by Electrospray Ionization Mass Spectrometry-Based Diffusion Measurements

• Published in: Analytical chemistry (Washington) Vol. 76; no. 5; pp. 1257 - 1263
• Main Authors: Clark, Sonya M, Konermann, Lars
• Format: Journal Article
• Language: English
• Published: Washington, DC American Chemical Society 01.03.2004
• Subjects: Analytical biochemistry: general aspects, technics, instrumentation; Analytical chemistry; Analytical, structural and metabolic biochemistry; Animals; Biological and medical sciences; Carbohydrate Metabolism; Carbohydrates - analysis; Carbohydrates - chemistry; Chemical bonds; Chemistry; Chickens; Diffusion; Exact sciences and technology; Fundamental and applied biological sciences. Psychology; Ligands; Mass spectrometry; Molecules; Muramidase - chemistry; Muramidase - metabolism; Protein Binding; Proteins; Spectrometric and optical methods; Spectrometry, Mass, Electrospray Ionization - methods; Time Factors; Enzyme; Ligand; Glucose; Maltose; Electrospray; Chemical ionization; Protein; Screening; Glycosidases; Hydrolases; Diffusion coefficient; O-Glycosidases; Molecular mass; Mass spectrometry; Lysozyme; Biological receptor
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac035230l

The application of a novel method for the identification of low-molecular-weight noncovalent ligands to a macromolecular target is reported. This technique is based on the measurement of analyte diffusion coefficients by electrospray mass spectrometry (ESI-MS) (Clark et al., Rapid Commun. Mass Spectrom. 2002, 16, 1454−1462). Potential ligands have large diffusion coefficients as long as they are free in solution. Binding to a macromolecular target, however, drastically reduces the diffusional mobility of any ligand species. Mixtures containing six different saccharides [ribose, rhamnose, glucose, maltose, maltotriose, and N,N‘,N‘ ‘-triacetylchitotriose (NAG3)] were screened for noncovalent binding to lysozyme. Of these six compounds, only NAG3 is known to bind to the protein. In “direct” binding tests, NAG3 shows a significantly reduced diffusion coefficient in the presence of the protein. No changes were observed for any of the other saccharides. In a second set of experiments, the use of a “competition” screening method was explored in which mixtures of candidate saccharides were tested for their ability to displace a reference ligand from the target. The addition of NAG3-containing mixtures significantly increased the diffusion coefficient of the reference ligand NAG4 (N,N‘,N‘ ‘,N‘ ‘‘-tetraacetylchitotetrose), whereas mixtures that did not contain NAG3 had no effect. These data clearly indicate the potential of ESI-MS-based diffusion measurements as a novel tool to screen compound libraries for binding to proteins and other macromolecular targets. In contrast to conventional ESI-MS-based ligand−receptor binding studies, this method does not rely on the preservation of noncovalent interactions in the gas phase.