Fibronectins are multifunctional glycoproteins that participate in blood coagulation, cell adhesion, differentiation and embryonic development. The basis of the multifunctional activities of fibronectins appears to reside in the multidomain structure of these molecules. We propose to use electron spin resonance (ESR) spin label methods to study the dynamic interactions between different regions of the fibronectin molecule. The transglutaminase crosslinking site and free sulfhydryl groups of the protein will be modified selectively with specific spin labels. The motions of fibronectin will be determined using both conventional and Saturation Transfer ESR methods. The effects of environmental conditions (e.g., pH, temperature and ionic strength) and interactions with other physiological substances (e.g., heparin, collagen, and fibrinogen) on the conformational states of fibronectins will be evaluated. Attempts will made to investigate the similarity and/or dissimilarity in solution properties between plasma and cellular fibronectins. In addition, the transglutaminase crosslinking site and free sulfhydryl groups of the protein will also be modified selectively with specific fluorescent labels. The changes in proximity relationship between different regions of the fibronectin molecule as affected by either environmental factors or interactions with other biomolecules will be determined by using fluorescence energy transfer methods. The studies should provide a deeper understanding of the three-dimensional relationships of various regions of fibronectin. Understanding the structure and dynamics of fibronectin underlies an interpretation of the vast array of physiological functions attributed to the protein. Finally, fibronectin is a prototype protein for other adhesive proteins such as laminin and chondronectin. Our long-term objective is to study the structure and function of these cell adhesive glycoproteins and to unravel the ways in which these proteins interact with the cell surface and the dynamic structure of extracellular matrices.
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