Tabtoxin production and resistance in P. syringae isolate BR2 is of considerable interest for numerous reasons. Molecular genetic munipulations are more amenable in Pseudomonads than most other monobactam producers. One of the ultimate goals of the research is to manipulate genes involved in antibiotic production (tabtoxin) and resistance. This ability would enable us to (a) increase yields of antibiotics, (b) make semi-synthetic antibiotics from pathway intermediates and possibly less toxic antibiotics (c) produce hybrid antibiotics due to interspecies transfer of antibiotic biosynthetic genes. Little information is available on the biosynthetic pathway in the producing organism. Monobactam, when isolated from nature exhibit weak antibiotic activity that may be modified by various chemical substitution to produce compounds that are more potent and resistant to B- lactamases. The widespread use of antibiotics has provided selective pressure for maintaining resistant organisms. Studies on antibiotic resistance have been useful to establish mechanisms of transfer of antibiotic resistances among bacteria. Second, these studies are useful for commercial strain improvement program since certain industries classical rely on the selection of antibiotic resistance as a way of isolating more efficient antibiotic producers. Tabtoxin production and resistance may serve as genetic markers for developing additional cloning vectors for the antibiotic-producing streptomycetes. The specific goal of the proposed research is to (1) determine the number and location of genes required for tabtoxin production and resistance (2) analyze gene products (3) determine the mechanism of spontaneous deletion that is required for tabtoxin production and resistance. The methods used include Southern analysis, DNA sequencing, Tn5 and site directed mutagenesis and isolation of toxin minus mutants.
