EFFECT OF THE SULFHYDRYL COMPOUND CYSTEAMINE ON GAMMA-RADIATION-INDUCED MUTATIONS IN DOUBLE-STRANDED M13 DNA

Sulfhydryl compounds can protect DNA against free-radical-induced DNA damages not only by scavenging of radicals, but also by chemical non-e nzymatic repair or modification of such damages by hydrogen-donation. To investigate the influence of chemical repair and modification on mu tations, induced by gamma-radiation-generated free radicals (. OH, . H ), phosphate-buffered aqueous solutions of double-stranded (ds) M13 DN A were exposed to gamma-rays under N-2 in the presence of 5 mM cysteam ine. The exposed DNA was subsequently transfected to wild-type E. coli and mutations in the mutational target were characterized. This targe t in fact contains three different target sequences, i.e., the lac pro moter/operator, the lacZ alpha gene and a 144 bp inframe insert. The m utation spectrum obtained was compared with those in the absence of cy steamine under N-2 and N2O. In the latter case, the ratio of . OH and . H available for reacting with DNA is about the same as under N-2 + c ysteamine. The results show that chemical repair and/or modification b y cysteamine of potentially lethal lesions takes place, leading to a m uch higher survival of ds M13 DNA in the presence of cysteamine than c ould be expected on basis of scavenging of . OH and . H alone. This hi gher survival appeared to be accompanied with a higher mutation induct ion. However, the N-2 + cysteamine mutation spectrum shows a remarkabl e resemblance with the N2O-spectrum. This holds for the total mutation target, as well as each of the three targets, although the mutations obtained in each of the three targets under the same irradiation condi tions are quite different. Thus, it can be concluded that cysteamine i s mainly effective on radiation-induced potentially lethal DNA lesions , and not so much on (pre)mutagenic damages. Moreover, the type of mut ation appeared to be strongly dependent on the mutational target seque nce.