An affinity selection-mass spectrometry method for the identification of small molecule ligands from self-encoded combinatorial libraries

Discovery of a novel antagonist of E. coli dihydrofolate reductase

D. Allen Annis, John Athanasopoulos, Patrick J. Curran, Jason S. Felsch, Krishna Kalghatgi, William H. Lee, Huw M. Nash, Jean Paul A. Orminati, Kristin E. Rosner, Gerald W. Shipps, G. R.A. Thaddupathy, Andrew N. Tyler, Lev Vilenchik, Carston R Wagner, Edward A. Wintner

Research output: Contribution to journalArticle

83 Citations (Scopus)

Abstract

The NeoGenesis Automated Ligand Identification System (ALIS), an affinity selection-mass spectrometry (AS-MS) process consisting of a rapid size-exclusion chromatography stage integrated with reverse-phase chromatography, electrospray mass spectrometry, and novel data searching algorithms, was used to screen mass-encoded, 2500-member combinatorial libraries, leading to the discovery of a novel, bioactive ligand for the anti-infective target Escherichia coli dihydrofolate reductase (DHFR). Synthesis of the mass-encoded, ligand-containing library, discussion of the deconvolution process for verifying the structure of the ligand through independent synthesis and screening in a small mixture (sub-library) format, and ALIS-MS/MS techniques to assign its regioisomeric connectivity are presented. ALIS-based competition experiments between the newly discovered ligand and other, known DHFR ligands, and biological activity assessments with stereo- and regioisomers of the hit compound confirm its DHFR-specific biological activity. The method described requires no foreknowledge of the structure or biochemistry of the protein target, consumes less than 1 μg protein to screen >2500 compounds in a single experiment, and enables screening of >250,000 compounds per system per day. These advantages highlight the potential of the ALIS method for drug discovery against genomic targets with unknown biological function, as well as validated targets for which traditional discovery efforts have failed.

Original languageEnglish (US)
Pages (from-to)77-83
Number of pages7
JournalInternational Journal of Mass Spectrometry
Volume238
Issue number2
DOIs
StatePublished - Nov 1 2004

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Tetrahydrofolate Dehydrogenase
Escherichia coli
Mass spectrometry
affinity
mass spectroscopy
Ligands
ligands
Molecules
system identification
molecules
Identification (control systems)
activity (biology)
chromatography
Bioactivity
Screening
screening
proteins
Proteins
Oxidoreductases
biochemistry

Keywords

  • Affinity selection-mass spectrometry
  • Combinatorial chemistry
  • Multidimensional chromatography

Cite this

An affinity selection-mass spectrometry method for the identification of small molecule ligands from self-encoded combinatorial libraries : Discovery of a novel antagonist of E. coli dihydrofolate reductase. / Annis, D. Allen; Athanasopoulos, John; Curran, Patrick J.; Felsch, Jason S.; Kalghatgi, Krishna; Lee, William H.; Nash, Huw M.; Orminati, Jean Paul A.; Rosner, Kristin E.; Shipps, Gerald W.; Thaddupathy, G. R.A.; Tyler, Andrew N.; Vilenchik, Lev; Wagner, Carston R; Wintner, Edward A.

In: International Journal of Mass Spectrometry, Vol. 238, No. 2, 01.11.2004, p. 77-83.

Research output: Contribution to journalArticle

Annis, DA, Athanasopoulos, J, Curran, PJ, Felsch, JS, Kalghatgi, K, Lee, WH, Nash, HM, Orminati, JPA, Rosner, KE, Shipps, GW, Thaddupathy, GRA, Tyler, AN, Vilenchik, L, Wagner, CR & Wintner, EA 2004, 'An affinity selection-mass spectrometry method for the identification of small molecule ligands from self-encoded combinatorial libraries: Discovery of a novel antagonist of E. coli dihydrofolate reductase', International Journal of Mass Spectrometry, vol. 238, no. 2, pp. 77-83. https://doi.org/10.1016/j.ijms.2003.11.022
Annis, D. Allen ; Athanasopoulos, John ; Curran, Patrick J. ; Felsch, Jason S. ; Kalghatgi, Krishna ; Lee, William H. ; Nash, Huw M. ; Orminati, Jean Paul A. ; Rosner, Kristin E. ; Shipps, Gerald W. ; Thaddupathy, G. R.A. ; Tyler, Andrew N. ; Vilenchik, Lev ; Wagner, Carston R ; Wintner, Edward A. / An affinity selection-mass spectrometry method for the identification of small molecule ligands from self-encoded combinatorial libraries : Discovery of a novel antagonist of E. coli dihydrofolate reductase. In: International Journal of Mass Spectrometry. 2004 ; Vol. 238, No. 2. pp. 77-83.
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