Measuring and modeling the kinetics of individual DNA-DNA polymerase complexes on a nanopore.

ABSTRACT: The assembly of a DNA-DNA polymerase binary complex is the precursory step in genome replication, in which the enzyme binds to the 3' junction created when a primer binds to its complementary substrate. In this study, we use an active control method for observing the binding interaction between Klenow fragment (exo-) (KF) in the bulk-phase chamber above an α-hemolysin (α-HL) nanopore and a single DNA molecule tethered noncovalently in the nanopore. Specifically, the control method regulates the temporal availability of the primer-template DNA to KF binding and unbinding above the nanopore, on millisecond-to-second time scales. Our nanopore measurements support a model that incorporates two mutually exclusive binding states of KF to DNA at the primer-template junction site, termed "weakly b

SUBMITTER: Wang H 

PROVIDER: S-EPMC3682681 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature