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Ligand for EPH-related kinase (LERK) 7 is the preferred high affinity ligand for the HEK receptor


Lackmann, M; Mann, RJ; Kravets, L; Smith, FM; Bucci, TA; Maxwell, KF; Howlett, GJ; Olsson, JE; Bos, TV; Cerretti, DP; Boyd, AW
1997-06-27
JOURNAL OF BIOLOGICAL CHEMISTRY
Journal Article
272
26
16521-16530
HEK is a member of the EPH-like receptor tyrosine kinase family, which appear to have roles in development and oncogenesis. Recently, we purified a soluble HEK ligand which is also a ligand (AL1) for the HEK-related receptor EHK1. Promiscuity appears to be a characteristic feature of interactions between the EPH-like receptors and their ligands, termed ligands for EPH-related kinases (LERKs). This prompted us to analyze the interactions between the HER exodomain and fusion proteins comprising candidate LERKs and the Fc portion of human IgG(1) (Fc) or a FLAG(TM)-peptide tag by surface plasmon resonance, size exclusion high performance liquid chromatography, sedimentation equilibrium, and transphosphorylation. Our results indicate that AL1/LERK7 is the preferred high-affinity ligand for HEK, forming a stable 1:1 complex with a dissociation constant of 12 nM. As expected the apparent affinities of bivalent fusion proteins of LERKs and the Fc portion of human IgG(1) had significantly reduced dissociation rates compared with their monovalent, FLAG(TM)-tagged derivatives. High-avidity binding of monovalent ligands can be achieved by antibody-mediated cross-linking of monovalent ligands and with LERK7 results in specific phosphorylation of the receptor. By extrapolation, our findings indicate that some of the reported LERK-receptor interactions are a consequence of the use of bivalent ligand or receptor constructs and may be functionally irrelevant.
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
PROTEIN-TYROSINE KINASE; SURFACE-PLASMON RESONANCE; BINDING CONSTANTS; MONOCLONAL-ANTIBODY; MOLECULAR-CLONING; KINETIC-ANALYSIS; GROWTH-FACTOR; CELL LINES; FAMILY; EXPRESSION
10.1074/jbc.272.26.16521
Refer to copyright notice on published article.

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Creation Date 1997-06-27 12:00:00