On the structure of the ends of lambda DNA

Abstract

The structure of the ends of a molecule of λ DNA has been deduced from the effect of DNA polymerase and exonuclease III on the infectivity of λ DNA and from the ability of the two ends of the molecule to cohere. The infectivity of λ DNA is destroyed when it primes DNA synthesis catalyzed by DNA polymerase. Inactivation depends on the presence of deoxynucleoside triphosphates and on the amount of enzyme added. The inactivating synthesis behaves like a repair of single-stranded regions, that is, their conversion to double strands, because it occurs at 15°C. Richardson, Inman & Kornberg (1964) found that DNA polymerase catalyzes repair but not net synthesis at temperatures lower than 20°C. Infectivity returns if the polymerase product is subsequently exposed to exonuclease III. The specificity of exonuclease III is such that it would be expected to remove nucleotides added by polymerase. This result shows that inactivation by polymerase is not due to degradation. In a complementary fashion the infectivity of native λ DNA is destroyed by exonuclease III and restored by subsequent synthesis catalyzed by DNA polymerase. The two ends of a molecule of λ DNA can cohere, specifically, to each other ( Hershey, Burgi & Ingraham, 1963 ; Ris & Chandler, 1963 ). To account for cohesiveness and for inactivation by DNA polymerase, it is proposed that 5′-hydroxyl (or 5′-phosphate)-terminated single strands protrude from both ends of the otherwise double-stranded molecule and that the two protruding single strands have complementary base sequences. Cohesion would result from base pairing between the protruding single strands. Synthesis primed by native λ DNA and catalyzed by DNA polymerase at 15°C would be expected to repair partially single-stranded ends and to destroy their cohesiveness. As the infectivity of λ DNA for helper-infected bacteria requires cohesiveness, polymerase would thus inactivate λ DNA. Treating the polymerase product with exonuclease III restores infectivity, according to this model, because removal of the added nucleotides restores cohesiveness.