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Isolation And Characterization Of Temperature-Sensitive Protein Synthesis Mutants Of Escherichia Coli By Directed Mutagenesis Of The Defective Bacteriophage Lambda Fus2, Kenton L. Lohman
Electronic Theses and Dissertations
Mutagenesis of the defective transducing bacteriophage lambda fus2 was used to isolate a collection of temperature-sensitive mutants of E. coli in the major ribosomal protein gene cluster. Four mutants were examined in detail. Two of the mutants were resistant to the ribosomal antibiotics neamine and spectinomycin. Another mutant was defective in 50S ribosomal subunit assembly at 42(DEGREES)C. The 30S subunit proteins S17 and S19 were changed in two different mutants. Each protein migrated as a more basic species in two-dimensional gels of ribosomal proteins. Ribosomes from each of the four mutants examined showed a temperature-dependent reduction in translational activity in …
Human Lung Tryptase Purification And Characterization, Timothy J. Smith
Human Lung Tryptase Purification And Characterization, Timothy J. Smith
Electronic Theses and Dissertations
Human lung tryptase (HLT), a mast cell derived trypsin-like enzyme, was isolated from whole human lung tissue obtained at autopsy. Increased yields from this purification process allowed extensive characterization of the enzyme. One of the critical steps in the purification scheme was the use of a linear heparin gradient to elute active material from cellulose phosphate. Gel filtration studies in 1.0 M NaCl yielded an apparent M(,r) of 135,000, and subsequent electrophoresis on sodium dodecyl sulfate-polyacrylamide gels demonstrated the presence of two active species with apparent M(,r) = 30,900 and 31,600. Enzymatic activity was sensitive to NaCl concentrations above 0.05 …