AzuR From the SmtB/ArsR Family of Transcriptional Repressors Regulates Metallothionein in Anabaena sp. Strain PCC 7120
Metallothioneins (MTs) are cysteine-rich, metal-sequestering cytosolic proteins that play a key role in maintaining metal homeostasis and detoxification. We had previously characterized NmtA, a MT from the heterocystous, nitrogen-fixing cyanobacterium sp. strain PCC 7120 and demonstrated its role in...
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Published in: | Frontiers in microbiology Vol. 12; p. 782363 |
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Main Authors: | , |
Format: | Journal Article |
Language: | English |
Published: |
Switzerland
Frontiers Media S.A
12-01-2022
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Subjects: | |
Online Access: | Get full text |
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Summary: | Metallothioneins (MTs) are cysteine-rich, metal-sequestering cytosolic proteins that play a key role in maintaining metal homeostasis and detoxification. We had previously characterized NmtA, a MT from the heterocystous, nitrogen-fixing cyanobacterium
sp. strain PCC 7120 and demonstrated its role in providing protection against cadmium toxicity. In this study, we illustrate the regulation of
NmtA by AzuR (Alr0831) belonging to the SmtB/ArsR family of transcriptional repressors. There is currently no experimental evidence for any functional role of AzuR. It is observed that
is located within the
operon but in the opposite orientation and remotely away from the
locus. Sequence analysis of AzuR revealed a high degree of sequence identity with
SmtB and a distinct α5 metal binding site similar to that of SmtB. In order to characterize AzuR, we overexpressed it in
and purified it by chitin affinity chromatography. Far-UV circular dichroism spectroscopy indicated that the recombinant AzuR protein possessed a properly folded structure. Glutaraldehyde cross-linking and size-exclusion chromatography revealed that AzuR exists as a dimer of ∼28 kDa in solution. Analysis of its putative promoter region [100 bp upstream of
open reading frame (ORF)] identified the presence of a 12-2-12 imperfect inverted repeat as the
-acting element important for repressor binding. Electrophoretic mobility shift assays (EMSAs) showed concentration-dependent binding of recombinant dimeric AzuR with the promoter indicating that NmtA is indeed a regulatory target of AzuR. Binding of AzuR to DNA was disrupted in the presence of metal ions like Zn
, Cd
, Cu
, Co
, Ni
, Pb
, and Mn
. The metal-dependent dissociation of protein-DNA complexes suggested the negative regulation of metal-inducible
expression by AzuR. Overexpression of
in its native strain
7120 enhanced the susceptibility to cadmium stress significantly. Overall, we propose a negative regulation of
MT by an α5 SmtB/ArsR metalloregulator AzuR. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Edited by: Rob Van Houdt, Belgian Nuclear Research Centre, Belgium Reviewed by: Vicente Mariscal, Institute of Plant Biochemistry and Photosynthesis, Spanish National Research Council (CSIC), Spain; Andrés González, Aragón Institute for Health Research (IIS Aragón), Spain This article was submitted to Antimicrobials, Resistance and Chemotherapy, a section of the journal Frontiers in Microbiology |
ISSN: | 1664-302X 1664-302X |
DOI: | 10.3389/fmicb.2021.782363 |