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Title: A semisynthetic strategy leads to alteration of the backbone amidate ligand in the NiSOD active site

Abstract

In this study, computational investigations have implicated the amidate ligand in nickel superoxide dismutase (NiSOD) in stabilizing Ni-centered redox catalysis and in preventing cysteine thiolate ligand oxidation. To test these predictions, we have used an experimental approach utilizing a semisynthetic scheme that employs native chemical ligation of a pentapeptide (HCDLP) to recombinant S. coelicolor NiSOD lacking these N-terminal residues, NΔ5-NiSOD. Wild-type enzyme produced in this manner exhibits the characteristic spectral properties of recombinant WT-NiSOD and is as catalytically active. The semisynthetic scheme was also employed to construct a variant where the amidate ligand was converted to a secondary amine, H1*-NiSOD, a novel strategy that retains a backbone N-donor atom. The H1*-NiSOD variant was found to have only ~1% of the catalytic activity of the recombinant wild-type enzyme, and had altered spectroscopic properties. X-ray absorption spectroscopy reveals a four-coordinate planar site with N2S2-donor ligands, consistent with electronic absorption spectroscopic results indicating that the Ni center in H1*-NiSOD is mostly reduced in the as-isolated sample, as opposed to 50:50 Ni(II)/Ni(III) mixture that is typical for the recombinant wild-type enzyme. The EPR spectrum of as-isolated H1*-NiSOD accounts for ~11% of the Ni in the sample and is similar to WT-NiSOD, but more axial,more » with gz < gx,y. 14N-hyperfine is observed on gz, confirming the addition of the apical histidine ligand in the Ni(III) complex. Lastly, the altered electronic properties and implications for redox catalysis are discussed in light of predictions based on synthetic and computational models.« less

Authors:
 [1];  [2];  [3];  [1];  [4];  [1]
  1. Univ. of Massachusetts, Amherst, MA (United States)
  2. Lex Company Research Lab., Shirley, MA (United States)
  3. New England Peptide, Gardner, MA (United States)
  4. Brookhaven National Lab. (BNL), Upton, NY (United States)
Publication Date:
Research Org.:
Brookhaven National Laboratory (BNL), Upton, NY (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES)
OSTI Identifier:
1224774
Alternate Identifier(s):
OSTI ID: 1335404
Report Number(s):
BNL-108507-2015-JA; BNL-111846-2016-JA
Journal ID: ISSN 0002-7863; R&D Project: CO004; KC0304030
Grant/Contract Number:  
SC00112704; SC0012704; AC02-98CH10886
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Journal of the American Chemical Society
Additional Journal Information:
Journal Volume: 137; Journal Issue: 28; Journal ID: ISSN 0002-7863
Publisher:
American Chemical Society (ACS)
Country of Publication:
United States
Language:
English
Subject:
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY

Citation Formats

Campeciño, Julius O., Dudycz, Lech W., Tumelty, David, Berg, Volker, Cabelli, Diane E., and Maroney, Michael J. A semisynthetic strategy leads to alteration of the backbone amidate ligand in the NiSOD active site. United States: N. p., 2015. Web. doi:10.1021/jacs.5b03629.
Campeciño, Julius O., Dudycz, Lech W., Tumelty, David, Berg, Volker, Cabelli, Diane E., & Maroney, Michael J. A semisynthetic strategy leads to alteration of the backbone amidate ligand in the NiSOD active site. United States. https://doi.org/10.1021/jacs.5b03629
Campeciño, Julius O., Dudycz, Lech W., Tumelty, David, Berg, Volker, Cabelli, Diane E., and Maroney, Michael J. 2015. "A semisynthetic strategy leads to alteration of the backbone amidate ligand in the NiSOD active site". United States. https://doi.org/10.1021/jacs.5b03629. https://www.osti.gov/servlets/purl/1224774.
@article{osti_1224774,
title = {A semisynthetic strategy leads to alteration of the backbone amidate ligand in the NiSOD active site},
author = {Campeciño, Julius O. and Dudycz, Lech W. and Tumelty, David and Berg, Volker and Cabelli, Diane E. and Maroney, Michael J.},
abstractNote = {In this study, computational investigations have implicated the amidate ligand in nickel superoxide dismutase (NiSOD) in stabilizing Ni-centered redox catalysis and in preventing cysteine thiolate ligand oxidation. To test these predictions, we have used an experimental approach utilizing a semisynthetic scheme that employs native chemical ligation of a pentapeptide (HCDLP) to recombinant S. coelicolor NiSOD lacking these N-terminal residues, NΔ5-NiSOD. Wild-type enzyme produced in this manner exhibits the characteristic spectral properties of recombinant WT-NiSOD and is as catalytically active. The semisynthetic scheme was also employed to construct a variant where the amidate ligand was converted to a secondary amine, H1*-NiSOD, a novel strategy that retains a backbone N-donor atom. The H1*-NiSOD variant was found to have only ~1% of the catalytic activity of the recombinant wild-type enzyme, and had altered spectroscopic properties. X-ray absorption spectroscopy reveals a four-coordinate planar site with N2S2-donor ligands, consistent with electronic absorption spectroscopic results indicating that the Ni center in H1*-NiSOD is mostly reduced in the as-isolated sample, as opposed to 50:50 Ni(II)/Ni(III) mixture that is typical for the recombinant wild-type enzyme. The EPR spectrum of as-isolated H1*-NiSOD accounts for ~11% of the Ni in the sample and is similar to WT-NiSOD, but more axial, with gz < gx,y. 14N-hyperfine is observed on gz, confirming the addition of the apical histidine ligand in the Ni(III) complex. Lastly, the altered electronic properties and implications for redox catalysis are discussed in light of predictions based on synthetic and computational models.},
doi = {10.1021/jacs.5b03629},
url = {https://www.osti.gov/biblio/1224774}, journal = {Journal of the American Chemical Society},
issn = {0002-7863},
number = 28,
volume = 137,
place = {United States},
year = {Wed Jul 01 00:00:00 EDT 2015},
month = {Wed Jul 01 00:00:00 EDT 2015}
}

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Cited by: 17 works
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Works referenced in this record:

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Works referencing / citing this record:

Oxidation State Dependent Coordination Modes: Accessing an Amidate-Supported Nickel(I) δ-bis(C−H) Agostic Complex
journal, October 2016


Oxidation State Dependent Coordination Modes: Accessing an Amidate-Supported Nickel(I) δ-bis(C−H) Agostic Complex
journal, October 2016


1,3-N,O-Complexes of late transition metals. Ligands with flexible bonding modes and reaction profiles
journal, January 2017


Engineered Metalloenzymes with Non‐Canonical Coordination Environments
journal, June 2018