A dual-constriction biological nanopore resolves homonucleotide sequences with high fidelity.
Nat Biotechnol
; 38(12): 1415-1420, 2020 12.
Article
em En
| MEDLINE
| ID: mdl-32632300
ABSTRACT
Single-molecule long-read DNA sequencing with biological nanopores is fast and high-throughput but suffers reduced accuracy in homonucleotide stretches. We now combine the CsgG nanopore with the 35-residue N-terminal region of its extracellular interaction partner CsgF to produce a dual-constriction pore with improved signal and base-calling accuracy for homopolymer regions. The electron cryo-microscopy structure of CsgG in complex with full-length CsgF shows that the 33 N-terminal residues of CsgF bind inside the ß-barrel of the pore, forming a defined second constriction. In complexes of CsgG bound to a 35-residue CsgF constriction peptide, the second constriction is separated from the primary constriction by ~25 Å. We find that both constrictions contribute to electrical signal modulation during single-stranded DNA translocation. DNA sequencing using a prototype CsgG-CsgF protein pore with two constrictions improved single-read accuracy by 25 to 70% in homopolymers up to 9 nucleotides long.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Nanoporos
/
Nucleotídeos
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article