This award provides funds to the Department of Biochemistry at Louisiana State University at Baton Rouge to purchase equipment that will help upgrade the core biochemistry lab course. This undergraduate Experimental Biochemistry course will focus on enzymology and molecular biology using a cloned full-length cDNA of Drosophila melanogaster alcohol dehydrogenase (ADH) to provide experimental materials. ADH will be purified from E. coli carrying the ADHcDNA cloned in an expression vector. The purified enzyme will be analyzed to determine the Km values for primary and secondary alcohols and the Ki values for competitive and non- competitive inhibitors. Purified ADH will be reversibly inactivated and reactivated through thio-disulfide interconversion. Effects of Nonsense, Missense and Deletion mutations will be examined by comparing ADH activity and antigenic response in the lysates of E. coli carrying the mutated ADHcDNAs. The ADH insert in M13 phage will be used to teach restriction mapping and DNA sequencing techniques. The principles of nucleic acid chemistry will be illustrated by determining the sequences of di- and tri- ribonucleotides with ribonuclease digestions, chromatography and UV spectra of the products under acidic, neutral and basic conditions. Denaturation of doubly-stranded DNA will be examined. Computer analysis and application are emphasized in appropriate experiments. The grantee is matching with award with non-Federal sources.
