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SCREENING METHOD FOR UBIQUITIN AND UBIQUITIN-LIKE PROTEIN SPECIFIC PROTEASES USING CI GENE OF LAMDA PHAGE, IN WHICH THE ENZYMES ARE USEFUL FOR DEVELOPMENT OF DRUGS FOR TREATING BRAIN NEURONAL DISEASES
SCREENING METHOD FOR UBIQUITIN AND UBIQUITIN-LIKE PROTEIN SPECIFIC PROTEASES USING CI GENE OF LAMDA PHAGE, IN WHICH THE ENZYMES ARE USEFUL FOR DEVELOPMENT OF DRUGS FOR TREATING BRAIN NEURONAL DISEASES
PURPOSE: A screening method for ubiquitin and ubiquitin-like protein specific proteases using cI gene of lambda phage is provided, thereby easily screening ubiquitin and ubiquitin-like protein specific proteases and simultaneously measuring activity of enzymes, so that the enzymes are useful for development of drugs for treating brain neuronal diseases. CONSTITUTION: The screening method for ubiquitin and ubiquitin-like protein specific proteases using cI gene of lambda phage comprises the steps of: (1) cloning Ub, UbI and cI genes, preparing a mutant of cI gene, preparing an expression plasmid of Ub-cI* or UbI-cI* fusion protein, and preparing transformed E. coli using the expression plasmid; (2) introducing cDNA library into the transformed E. coli using recombinant lambda-phage; (3) selecting E. coli expressing desired protein by heat shock at 42 deg. C for 2 hours; and analyzing enzyme activity of the expressed protein, wherein the Ub-cI* or UbI-cI* fusion protein is produced by fusing ubiquitin-like protein Nedd8 or SUMO-1 with cI protein mutant(cI*); and the fusion protein expression plasmid is pACYC184.
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