PROTEIN MOLECULAR BIOLOGY OF TISSUE FACTOR

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Tissue factor (TF) is the primary and pivotal molecule responsible for initiation of the coagulation serine protease cascades. Expressed as a cell surface transmembrane receptor it initiates the assembly of the binary complex TF VIIa and guides the efficient assembly of the ternary complex that produces the active serine proteases Xa and IXa. We will address in detail the structural basis for TF function through model- guided site-directed mutagenesis coupled with integration with three dimensional structural data to elucidate the protein molecular biology of TF and the basis for the function of this receptor. The goals will be to: (i) define the critical residues and characterize functional replacements to develop structure to function interpretations of the molecular basis for the recognition and high affinity binding of VII and VIIa; (ii) investigate the structural basis of substrate X and IX coordination and specific proteolytic activation; (iii) explore the structural role of TF in the activation of VII; (iv) explore the role of flexibility and spacing above the cell and phospholipid membrane in regulation of the efficiency of the TF regulated initiation complex; (v) explore the role of TF dimerization in regulation of the efficiency of the initiation complex. This information will be necessary to interpret the three dimensional data from crystallography and high resolution NMR. In addition, the fundamental properties of TF and the assignment of classes of function to substructure will provide a detailed analysis of the structural biology of a model of the family of Cytokine/Hematopoietic Growth Factor Receptors.