Nucleotide selection by the Y-family DNA polymerase Dpo4 involves template translocation and misalignment
Author(s)
Brenlla, A
Markiewicz, RP
Rueda, D
Romano, LJ
Type
Journal Article
Abstract
Y-family DNA polymerases play a crucial role in translesion DNA synthesis. Here, we have characterized the binding kinetics and conformational dynamics of the Y-family polymerase Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) using single-molecule fluorescence. We find that in the absence of dNTPs, the binary complex shuttles between two different conformations within ∼1 s. These data are consistent with prior crystal structures in which the nucleotide binding site is either occupied by the terminal base pair (preinsertion conformation) or empty following Dpo4 translocation by 1 base pair (insertion conformation). Most interestingly, on dNTP binding, only the insertion conformation is observed and the correct dNTP stabilizes this complex compared with the binary complex, whereas incorrect dNTPs destabilize it. However, if the n+1 template base is complementary to the incoming dNTP, a structure consistent with a misaligned template conformation is observed, in which the template base at the n position loops out. This structure provides evidence for a Dpo4 mutagenesis pathway involving a transient misalignment mechanism.
Date Issued
2014-02-01
Online Publication Date
2014-02-01
2021-05-04T09:35:00Z
Date Acceptance
2013-10-25
ISSN
0305-1048
Publisher
Oxford University Press
Start Page
2555
End Page
2563
Journal / Book Title
Nucleic Acids Research
Volume
42
Issue
4
Copyright Statement
© The Author(s) 2013. Published by Oxford University Press.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
Source Database
web-of-science
Identifier
https://academic.oup.com/nar/article/42/4/2555/2435430
http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000332381000046&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=1ba7043ffcc86c417c072aa74d649202
Subjects
Science & Technology
Life Sciences & Biomedicine
Biochemistry & Molecular Biology
RESONANCE ENERGY-TRANSFER
SINGLE-MOLECULE FRET
I KLENOW FRAGMENT
CONFORMATIONAL-CHANGES
STRUCTURAL DIVERSITY
CRYSTAL-STRUCTURE
DAMAGE REPAIR
HIGH-FIDELITY
ACTIVE-SITE
ERROR-PRONE
DNA Polymerase beta
DNA Primers
Deoxyribonucleotides
Protein Conformation
Protein Transport
Sulfolobus solfataricus
Templates, Genetic
Sulfolobus solfataricus
DNA Polymerase beta
Deoxyribonucleotides
DNA Primers
Protein Conformation
Protein Transport
Templates, Genetic
Science & Technology
Life Sciences & Biomedicine
Biochemistry & Molecular Biology
RESONANCE ENERGY-TRANSFER
SINGLE-MOLECULE FRET
I KLENOW FRAGMENT
CONFORMATIONAL-CHANGES
STRUCTURAL DIVERSITY
CRYSTAL-STRUCTURE
DAMAGE REPAIR
HIGH-FIDELITY
ACTIVE-SITE
ERROR-PRONE
Developmental Biology
05 Environmental Sciences
06 Biological Sciences
08 Information and Computing Sciences
Publication Status
Published
Date Publish Online
2013-11-21