Methods for generating phosphorylation site-specific immunological reagents

Anderson, Carl W; Appella, Ettore; Sakaguchi, Kazuyasu

Methods for generating phosphorylation site-specific immunological reagents

Patent · Sun Dec 31 23:00:00 EST 2000

OSTI ID:874090

Anderson, Carl W ^[1]; Appella, Ettore ^[2]; Sakaguchi, Kazuyasu ^[2]

Stony Brook, NY
Montgomery, MD

The present invention provides methods for generating phosphorylation site-specific immunological reagents. More specifically, a phosphopeptide mimetic is incorporated into a polypeptide in place of a phosphorylated amino acid. The polypeptide is used as antigen by standard methods to generate either monoclonal or polyclonal antibodies which cross-react with the naturally phosphorylated polypeptide. The phosphopeptide mimetic preferably contains a non-hydrolyzable linkage from the appropriate carbon atom of the amino acid residue to a phosphate group. A preferred linkage is a CF.sub.2 group. Such a linkage is used to generate the phosphoserine mimetic F.sub.2 Pab, which is incorporated into a polypeptide sequence derived from p53 to produce antibodies which recognize a specific phosphorylation state of p53. A CF.sub.2 group linkage is also used to produce the phosphothreonine mimetic F.sub.2 Pmb, and to produce the phosphotyrosine mimetic, F.sub.2 Pmp.

Research Organization:: Brookhaven National Laboratory (BNL), Upton, NY

DOE Contract Number:: AC02-98CH10886

Assignee:: Brookhaven Science Associates (Upton, NY)

Patent Number(s):: US 6309863

OSTI ID:: 874090

Country of Publication:: United States

Language:: English

References (11)

Synthesis and application of N-Boc-L-2-amino-4-(diethylphosphono)-4,4-difluorobutanoic acid for solid-phase synthesis of nonhydrolyzable phosphoserine peptide analogues Otaka, Akira; Miyoshi, Kengo; Burke, Terrence R. Tetrahedron Letters, Vol. 36, Issue 6 https://doi.org/10.1016/0040-4039(94)02374-K	journal	February 1995
Regulation of CREB phosphorylation in the suprachiasmatic nucleus by light and a circadian clock Ginty, D.; Kornhauser, J.; Thompson, M. Science, Vol. 260, Issue 5105 https://doi.org/10.1126/science.8097062	journal	April 1993
DNA Damage-Induced Phosphorylation of p53 Alleviates Inhibition by MDM2 Shieh, Sheau-Yann; Ikeda, Masako; Taya, Yoichi Cell, Vol. 91, Issue 3 https://doi.org/10.1016/S0092-8674(00)80416-X	journal	October 1997
Production and Characterization of a Novel Monoclonal Antibody Against Phosphorylated T Cell Receptor Zeta Chain Conrad, P. J.; Seng, T.; Barbosa, J. Hybridoma, Vol. 16, Issue 2 https://doi.org/10.1089/hyb.1997.16.167	journal	April 1997
Rho-Kinase Phosphorylates COOH-terminal Threonines of Ezrin/Radixin/Moesin (ERM) Proteins and Regulates Their Head-to-Tail Association Matsui, Takeshi; Maeda, Masato; Doi, Yoshinori Journal of Cell Biology, Vol. 140, Issue 3 https://doi.org/10.1083/jcb.140.3.647	journal	February 1998
Why Is Phosphonodifluoromethyl Phenylalanine a More Potent Inhibitory Moiety Than Phosphonomethyl Phenylalanine Toward Protein-Tyrosine Phosphatases Chen, L.; Wu, L.; Otaka, A. Biochemical and Biophysical Research Communications, Vol. 216, Issue 3 https://doi.org/10.1006/bbrc.1995.2716	journal	November 1995
DNA damage activates p53 through a phosphorylation-acetylation cascade Sakaguchi, K.; Herrera, J. E.; Saito, S. 'i. Genes & Development, Vol. 12, Issue 18 https://doi.org/10.1101/gad.12.18.2831	journal	September 1998
DNA damage induces phosphorylation of the amino terminus of p53 Siliciano, J. D.; Canman, C. E.; Taya, Y. Genes & Development, Vol. 11, Issue 24 https://doi.org/10.1101/gad.11.24.3471	journal	December 1997
Nerve growth factor activates a Ras-dependent protein kinase that stimulates c-fos transcription via phosphorylation of CREB Ginty, David D.; Bonni, Azad; Greenberg, Michael E. Cell, Vol. 77, Issue 5 https://doi.org/10.1016/0092-8674(94)90055-8	journal	June 1994
Specific Localization of Serine 19 Phosphorylated Myosin II during Cell Locomotion and Mitosis of Cultured Cells Matsumura, Fumio; Ono, Shoichiro; Yamakita, Yoshihiko Journal of Cell Biology, Vol. 140, Issue 1 https://doi.org/10.1083/jcb.140.1.119	journal	January 1998
Potent Inhibition of Insulin Receptor Dephosphorylation by a Hexamer Peptide Containing the Phosphotyrosyl Mimetic F2Pmp Burke, T. R.; Kole, H. K.; Roller, P. P. Biochemical and Biophysical Research Communications, Vol. 204, Issue 1 https://doi.org/10.1006/bbrc.1994.2435	journal	October 1994

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Methods for generating phosphorylation site-specific immunological reagents

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References (11)

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