ACTIVE CONFORMATION OF LAMININ PEPTIDE 11

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The 67 kDa high affinity laminin binding protein (LBP) is a cell surface protein whose expression shows a positive correlation with progression in many solid tumors. This correlation likely results from the ability of the protein to mediate high affinity interactions of a metastatic tumor cells with basement membrane laminin molecules. The major ligand binding domain for the LBP occurs in the laminin-1 b chain at the COPGYIGSR (peptide 11) site. The investigators have preliminary data (from NMR Tr-NOESY experiments) on the conformation of peptide 11 when bound to isolated 67 kDa LBP, but experiments have been restricted by the low amount of available LBP. The investigators can not produce sufficient protein from their chinese hamster ovary (CHO) overexpression system for routine NMR experiments, and, in the present proposal, they plan to complete NMR structural studies on peptide 11 bound to the LBP. The investigators will also pursue structural studies on promising cyclic analogs of peptide 11. The peptides, themselves, have very short lifetimes in the blood stream and modest affinity, whereas non-peptide structural mimetics are expected to have significantly improved pharmacological characteristics. Such mimetics could provide extremely useful adjuvant therapies for malignant solid tumors. The long range goal of the work is to use the active conformation of peptide 11, when it is bound to the 67 kDa LBP, as a template for structure-based design of novel anti- metastatic drugs. This, however, requires the development of a very accurate template structure which the studies outlined in the current application would produce. The investigators will also undertake a series of experiments to increase our knowledge of the biochemistry and mechanism of action of the membrane associated form of the LBP. These experiments will use our peptide 11 based photolabelling probe to facilitate identification of the LBP residues which interact with peptide 11. Other experiments will evaluate the postulate that the 67 kDa LBP may act in concert with the a6b1 integrin during tissue invasion.