Skip to main content

Lund University Publications

LUND UNIVERSITY LIBRARIES

Nucleotide binding to the ATP-cone in anaerobic ribonucleotide reductases allosterically regulates activity by modulating substrate binding

Bimai, Ornella ; Banerjee, Ipsita LU ; Grinberg, Inna Rozman ; Huang, Ping LU ; Hultgren, Lucas LU ; Ekström, Simon LU ; Lundin, Daniel ; Sjöberg, Britt-Marie and Logan, Derek T. LU orcid (2024) In eLife 12.
Abstract

A small, nucleotide-binding domain, the ATP-cone, is found at the N-terminus of most ribonucleotide reductase (RNR) catalytic subunits. By binding adenosine triphosphate (ATP) or deoxyadenosine triphosphate (dATP) it regulates the enzyme activity of all classes of RNR. Functional and structural work on aerobic RNRs has revealed a plethora of ways in which dATP inhibits activity by inducing oligomerisation and preventing a productive radical transfer from one subunit to the active site in the other. Anaerobic RNRs, on the other hand, store a stable glycyl radical next to the active site and the basis for their dATP-dependent inhibition is completely unknown. We present biochemical, biophysical, and structural information on the effects... (More)

A small, nucleotide-binding domain, the ATP-cone, is found at the N-terminus of most ribonucleotide reductase (RNR) catalytic subunits. By binding adenosine triphosphate (ATP) or deoxyadenosine triphosphate (dATP) it regulates the enzyme activity of all classes of RNR. Functional and structural work on aerobic RNRs has revealed a plethora of ways in which dATP inhibits activity by inducing oligomerisation and preventing a productive radical transfer from one subunit to the active site in the other. Anaerobic RNRs, on the other hand, store a stable glycyl radical next to the active site and the basis for their dATP-dependent inhibition is completely unknown. We present biochemical, biophysical, and structural information on the effects of ATP and dATP binding to the anaerobic RNR from Prevotella copri. The enzyme exists in a dimer– tetramer equilibrium biased towards dimers when two ATP molecules are bound to the ATP-cone and tetramers when two dATP molecules are bound. In the presence of ATP, P. copri NrdD is active and has a fully ordered glycyl radical domain (GRD) in one monomer of the dimer. Binding of dATP to the ATP-cone results in loss of activity and increased dynamics of the GRD, such that it cannot be detected in the cryo-EM structures. The glycyl radical is formed even in the dATP-bound form, but the substrate does not bind. The structures implicate a complex network of interactions in activity regulation that involve the GRD more than 30 Å away from the dATP molecules, the allosteric substrate specificity site and a conserved but previously unseen flap over the active site. Taken together, the results suggest that dATP inhibition in anaerobic RNRs acts by increasing the flexibility of the flap and GRD, thereby preventing both substrate binding and radical mobilisation.

(Less)
Please use this url to cite or link to this publication:
author
; ; ; ; ; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
eLife
volume
12
article number
RP89292
pages
35 pages
publisher
eLife Sciences Publications
external identifiers
  • scopus:85197748338
  • pmid:38968292
ISSN
2050-084X
DOI
10.7554/ELIFE.89292
language
English
LU publication?
yes
additional info
Publisher Copyright: © Bimai et al.
id
bee7ab88-2e7a-4920-bf3f-f931cfa795f3
date added to LUP
2024-08-12 09:13:12
date last changed
2024-08-14 11:42:40
@article{bee7ab88-2e7a-4920-bf3f-f931cfa795f3,
  abstract     = {{<p>A small, nucleotide-binding domain, the ATP-cone, is found at the N-terminus of most ribonucleotide reductase (RNR) catalytic subunits. By binding adenosine triphosphate (ATP) or deoxyadenosine triphosphate (dATP) it regulates the enzyme activity of all classes of RNR. Functional and structural work on aerobic RNRs has revealed a plethora of ways in which dATP inhibits activity by inducing oligomerisation and preventing a productive radical transfer from one subunit to the active site in the other. Anaerobic RNRs, on the other hand, store a stable glycyl radical next to the active site and the basis for their dATP-dependent inhibition is completely unknown. We present biochemical, biophysical, and structural information on the effects of ATP and dATP binding to the anaerobic RNR from Prevotella copri. The enzyme exists in a dimer– tetramer equilibrium biased towards dimers when two ATP molecules are bound to the ATP-cone and tetramers when two dATP molecules are bound. In the presence of ATP, P. copri NrdD is active and has a fully ordered glycyl radical domain (GRD) in one monomer of the dimer. Binding of dATP to the ATP-cone results in loss of activity and increased dynamics of the GRD, such that it cannot be detected in the cryo-EM structures. The glycyl radical is formed even in the dATP-bound form, but the substrate does not bind. The structures implicate a complex network of interactions in activity regulation that involve the GRD more than 30 Å away from the dATP molecules, the allosteric substrate specificity site and a conserved but previously unseen flap over the active site. Taken together, the results suggest that dATP inhibition in anaerobic RNRs acts by increasing the flexibility of the flap and GRD, thereby preventing both substrate binding and radical mobilisation.</p>}},
  author       = {{Bimai, Ornella and Banerjee, Ipsita and Grinberg, Inna Rozman and Huang, Ping and Hultgren, Lucas and Ekström, Simon and Lundin, Daniel and Sjöberg, Britt-Marie and Logan, Derek T.}},
  issn         = {{2050-084X}},
  language     = {{eng}},
  month        = {{07}},
  publisher    = {{eLife Sciences Publications}},
  series       = {{eLife}},
  title        = {{Nucleotide binding to the ATP-cone in anaerobic ribonucleotide reductases allosterically regulates activity by modulating substrate binding}},
  url          = {{http://dx.doi.org/10.7554/ELIFE.89292}},
  doi          = {{10.7554/ELIFE.89292}},
  volume       = {{12}},
  year         = {{2024}},
}