This application proposes studies directed towards the identification and characterization of novel brain specific proteins. Rat brain cDNA clones were selected which correspond to mRNAs expressed in brain but not in liver or kidney. The nucleotide sequence of one such clone (plB236) has been determined, providing the amino acid sequence of the protein (1B236) encoded by the cloned mRNA. Antibodies raised against synthetic peptides corresponding to regions of this sequence detect immunoreactivity in a series of widely distributed neuronal structures in rat brain distinct from any other immunocytochemically defined neuronal system. In addition, the amino acid sequence of 1B236 has structural features in common with neuropeptide precursors, suggesting that the 1B236 protein may be cleaved to generate several novel neuropeptides. Preliminary evidence indicates that 1B236 immunoreactivity can be detected in rat brain extracts as both a 100,000 dalton glycoprotein and as processed peptides (15-40 amino acids). Synthetic peptides corresponding to 1B236 sequences have behavioral and electrophysiological activity. It is the aim of the proposed studies to: 1. Determine the predominant molecular forms of 1B236 immunoreactivity in rat brain and their relative amounts. 2. Isolate and characterize the 1B236 protein from rat brain. 3. Determine the exact forms of any peptides generated from the 1B236 protein in vivo and released in vitro from nerve terminals. 4. Deduce the sequence of steps involved in the post-translational processing of the 1B236 protein. 5. Determine the structure of the gene coding for the 1B236 protein. 6. Investigate the expression of the 1B236 protein and its products during development of the rat. 7. Test for similar molecules in other species. These studies will generate structural, genetic and developmental information on a novel brain specific protein which may represent a previously undetected neurotransmitter system. The data will provide the basis for functional testing of this protein and its peptide products and may provide a new avenue for the elucidation of neurologic disorders.
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