TY - JOUR
T1 - An affinity selection-mass spectrometry method for the identification of small molecule ligands from self-encoded combinatorial libraries
T2 - Discovery of a novel antagonist of E. coli dihydrofolate reductase
AU - Annis, D. Allen
AU - Athanasopoulos, John
AU - Curran, Patrick J.
AU - Felsch, Jason S.
AU - Kalghatgi, Krishna
AU - Lee, William H.
AU - Nash, Huw M.
AU - Orminati, Jean Paul A.
AU - Rosner, Kristin E.
AU - Shipps, Gerald W.
AU - Thaddupathy, G. R.A.
AU - Tyler, Andrew N.
AU - Vilenchik, Lev
AU - Wagner, Carston R
AU - Wintner, Edward A.
PY - 2004/11/1
Y1 - 2004/11/1
N2 - 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.
AB - 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.
KW - Affinity selection-mass spectrometry
KW - Combinatorial chemistry
KW - Multidimensional chromatography
UR - http://www.scopus.com/inward/record.url?scp=8444221197&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=8444221197&partnerID=8YFLogxK
U2 - 10.1016/j.ijms.2003.11.022
DO - 10.1016/j.ijms.2003.11.022
M3 - Article
AN - SCOPUS:8444221197
SN - 1387-3806
VL - 238
SP - 77
EP - 83
JO - International Journal of Mass Spectrometry
JF - International Journal of Mass Spectrometry
IS - 2
ER -