The binding of near-ultraviolet light-induced tryptophan photoproduct(s) to DNA

Abstract

To test one possible mode of toxicity of l-tryptophan photoproduct(s) to bacterial cells, we have examined the binding of near-ultraviolet light-treated tryptophan to purified Escherichia coli DNA in vitro. The results show that co-irradiated (or pre-irradiated) [ 3H]tryptophan binds to purified DNA as assayed by trichloroacetic acid co-precipitation of DNA and 3H counts on cellulose filters. This was supported by co-sedimentation of DNA and 3H photoproduct(s) on CsCl gradients. Hot trichloroacetic acid extraction or enzymatic digestion of DNA prevents filter binding. The binding is most efficient when tryptophan and DNA are co-irradiated. Under these conditions, binding is more efficient with denatured rather than native DNA. From kinetic studies, the binding is DNA-dependent at constant doses of near-ultraviolet light. At a dose of 2.16 · 10 6 ergs · mm −2 we estimate that 100–500 l-[ 3H]tryptophan equivalents are bound per E. coli genome equivalent. The binding does not occur with another aromatic amino acid such as tyrosine.