Abstract
A precise and rapid method for identifying sites of interaction between proteins was demonstrated; the basis of the method is direct mass spectrometric readout from the complex to determine the specific components of the proteins that interact--a method termed affinity-directed mass spectrometry. The strategy was used to define the region of interaction of a protein growth factor with a monoclonal antibody. A combination of proteolytic digestion and affinity-directed mass spectrometry was used to rapidly determine the approximate location of a continuous binding epitope within the growth factor. The precise boundaries of the binding epitope were determined by affinity-directed mass spectrometric analysis of sets of synthetic peptide ladders that span the approximate binding region. In addition to the mapping of such linear epitopes, affinity-directed mass spectrometry can be applied to the mapping of other types of molecule-molecule contacts, including ligand-receptor and protein-oligonucleotide interactions.
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Selected References
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- Abraham J. A., Whang J. L., Tumolo A., Mergia A., Friedman J., Gospodarowicz D., Fiddes J. C. Human basic fibroblast growth factor: nucleotide sequence and genomic organization. EMBO J. 1986 Oct;5(10):2523–2528. doi: 10.1002/j.1460-2075.1986.tb04530.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Beavis R. C., Chait B. T. Rapid, sensitive analysis of protein mixtures by mass spectrometry. Proc Natl Acad Sci U S A. 1990 Sep;87(17):6873–6877. doi: 10.1073/pnas.87.17.6873. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chait B. T., Kent S. B. Weighing naked proteins: practical, high-accuracy mass measurement of peptides and proteins. Science. 1992 Sep 25;257(5078):1885–1894. doi: 10.1126/science.1411504. [DOI] [PubMed] [Google Scholar]
- Chait B. T., Wang R., Beavis R. C., Kent S. B. Protein ladder sequencing. Science. 1993 Oct 1;262(5130):89–92. doi: 10.1126/science.8211132. [DOI] [PubMed] [Google Scholar]
- Cohen S. L., Ferré-D'Amaré A. R., Burley S. K., Chait B. T. Probing the solution structure of the DNA-binding protein Max by a combination of proteolysis and mass spectrometry. Protein Sci. 1995 Jun;4(6):1088–1099. doi: 10.1002/pro.5560040607. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Covey T. R., Huang E. C., Henion J. D. Structural characterization of protein tryptic peptides via liquid chromatography/mass spectrometry and collision-induced dissociation of their doubly charged molecular ions. Anal Chem. 1991 Jul 1;63(13):1193–1200. doi: 10.1021/ac00013a003. [DOI] [PubMed] [Google Scholar]
- Cunningham B. C., Jhurani P., Ng P., Wells J. A. Receptor and antibody epitopes in human growth hormone identified by homolog-scanning mutagenesis. Science. 1989 Mar 10;243(4896):1330–1336. doi: 10.1126/science.2466339. [DOI] [PubMed] [Google Scholar]
- Darnell J. E., Jr, Kerr I. M., Stark G. R. Jak-STAT pathways and transcriptional activation in response to IFNs and other extracellular signaling proteins. Science. 1994 Jun 3;264(5164):1415–1421. doi: 10.1126/science.8197455. [DOI] [PubMed] [Google Scholar]
- Devlin J. J., Panganiban L. C., Devlin P. E. Random peptide libraries: a source of specific protein binding molecules. Science. 1990 Jul 27;249(4967):404–406. doi: 10.1126/science.2143033. [DOI] [PubMed] [Google Scholar]
- Fairman R., Beran-Steed R. K., Anthony-Cahill S. J., Lear J. D., Stafford W. F., 3rd, DeGrado W. F., Benfield P. A., Brenner S. L. Multiple oligomeric states regulate the DNA binding of helix-loop-helix peptides. Proc Natl Acad Sci U S A. 1993 Nov 15;90(22):10429–10433. doi: 10.1073/pnas.90.22.10429. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Geysen H. M., Meloen R. H., Barteling S. J. Use of peptide synthesis to probe viral antigens for epitopes to a resolution of a single amino acid. Proc Natl Acad Sci U S A. 1984 Jul;81(13):3998–4002. doi: 10.1073/pnas.81.13.3998. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Houghten R. A., Pinilla C., Blondelle S. E., Appel J. R., Dooley C. T., Cuervo J. H. Generation and use of synthetic peptide combinatorial libraries for basic research and drug discovery. Nature. 1991 Nov 7;354(6348):84–86. doi: 10.1038/354084a0. [DOI] [PubMed] [Google Scholar]
- Kurokawa T., Sasada R., Iwane M., Igarashi K. Cloning and expression of cDNA encoding human basic fibroblast growth factor. FEBS Lett. 1987 Mar 9;213(1):189–194. doi: 10.1016/0014-5793(87)81489-8. [DOI] [PubMed] [Google Scholar]
- Lam K. S., Salmon S. E., Hersh E. M., Hruby V. J., Kazmierski W. M., Knapp R. J. A new type of synthetic peptide library for identifying ligand-binding activity. Nature. 1991 Nov 7;354(6348):82–84. doi: 10.1038/354082a0. [DOI] [PubMed] [Google Scholar]
- Laver W. G., Air G. M., Webster R. G., Smith-Gill S. J. Epitopes on protein antigens: misconceptions and realities. Cell. 1990 May 18;61(4):553–556. doi: 10.1016/0092-8674(90)90464-p. [DOI] [PubMed] [Google Scholar]
- Mitchell P. J., Tjian R. Transcriptional regulation in mammalian cells by sequence-specific DNA binding proteins. Science. 1989 Jul 28;245(4916):371–378. doi: 10.1126/science.2667136. [DOI] [PubMed] [Google Scholar]
- Nelson R. W., Krone J. R., Bieber A. L., Williams P. Mass spectrometric immunoassay. Anal Chem. 1995 Apr 1;67(7):1153–1158. doi: 10.1021/ac00103a003. [DOI] [PubMed] [Google Scholar]
- Parmley S. F., Smith G. P. Antibody-selectable filamentous fd phage vectors: affinity purification of target genes. Gene. 1988 Dec 20;73(2):305–318. doi: 10.1016/0378-1119(88)90495-7. [DOI] [PubMed] [Google Scholar]
- Schnölzer M., Alewood P., Jones A., Alewood D., Kent S. B. In situ neutralization in Boc-chemistry solid phase peptide synthesis. Rapid, high yield assembly of difficult sequences. Int J Pept Protein Res. 1992 Sep-Oct;40(3-4):180–193. doi: 10.1111/j.1399-3011.1992.tb00291.x. [DOI] [PubMed] [Google Scholar]
- Scott J. K., Smith G. P. Searching for peptide ligands with an epitope library. Science. 1990 Jul 27;249(4967):386–390. doi: 10.1126/science.1696028. [DOI] [PubMed] [Google Scholar]
- Suckau D., Köhl J., Karwath G., Schneider K., Casaretto M., Bitter-Suermann D., Przybylski M. Molecular epitope identification by limited proteolysis of an immobilized antigen-antibody complex and mass spectrometric peptide mapping. Proc Natl Acad Sci U S A. 1990 Dec;87(24):9848–9852. doi: 10.1073/pnas.87.24.9848. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zhao Y., Chalt B. T. Protein epitope mapping by mass spectrometry. Anal Chem. 1994 Nov 1;66(21):3723–3726. doi: 10.1021/ac00093a029. [DOI] [PubMed] [Google Scholar]