The long-term objective of this project is to analyze the elements of structure in a multifunctional, structural protein which give rise to the different functions that this protein performs. The model system being used is a procaryotic virus assembly protein which is required for virus adsorption and nucleic acid injection. The main tool to be used will be genetics. One of the most interesting functions of this protein is its assembly function in viral morphogenesis. How do structural proteins combine in tight non-covalent associations with their target protein(s)? This proposal focuses on this aspect; that is, what are the molecular interactions and mechanisms used in supramolecular structure formation? Because structure is a common attribute of all living things, understanding the mechanisms and regulation of its formation is basic knowledge applicable to developmental biology and the study of birth defects. A detailed analysis of several systems which are amenable to intense study should provide some understanding of how normal structures are formed; and, what can go wrong during dismorphogenesis.
The specific aims of this proposal are: to subject this model protein to an intense mutational analysis using both conventional and """"""""reverse"""""""" genetic techniques which have spawned from the recombinant DNA technologies; to identify the functions of this structural protein which are lost or impaired by these genetic lesions, to locate and identify at the nucleotide level where these mutations have occurred, to derive from this information the domain organization of this protein both in functional and structural terms and to identify those elements of structure which are involved in its assembly activity. Mutagenesis of this protein will be accomplished both by standard phage genetics and mutagenesis of the cloned gene. Both the isolation and the characterization of these mutations will use the tools of modern recombinant DNA technology and gene fusion. Functional analysis of the mutations will be done through established assays for this protein's activities. Identification of the mutational events will be done through DNA sequencing.
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