Sequence-dependent mechanochemical coupling of helicase translocation and unwinding at single-nucleotide resolution.

ABSTRACT: We used single-molecule picometer-resolution nanopore tweezers (SPRNT) to resolve the millisecond single-nucleotide steps of superfamily 1 helicase PcrA as it translocates on, or unwinds, several kilobase-long DNA molecules. We recorded more than two million enzyme steps under various assisting and opposing forces in diverse adenosine tri- and diphosphate conditions to comprehensively explore the mechanochemistry of PcrA motion. Forces applied in SPRNT mimic forces and physical barriers PcrA experiences in vivo, such as when the helicase encounters bound proteins or duplex DNA. We show how PcrA's kinetics change with such stimuli. SPRNT allows for direct association of the underlying DNA sequence with observed enzyme kinetics. Our data reveal that the underlying DNA sequence passing

SUBMITTER: Laszlo AH 

PROVIDER: S-EPMC9457475 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature