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The activity of barley NADPH-dependent thioredoxin reductase C is independent of the oligomeric state of the protein: tetrameric structure determined by cryo-electron microscopy

Peterson Wulff, Ragna LU ; Lundqvist, J.; Rutsdottir, Gudrun LU ; Hansson, A.; Stenbaek, A.; Elmlund, D.; Elmlund, H.; Jensen, P. E. and Hansson, Mats LU (2011) In Biochemistry 50(18). p.3713-3723
Abstract
Thioredoxin and thioredoxin reductase can regulate cell metabolism through redox regulation of disulfide bridges or through removal of H(2)O(2). These two enzymatic functions are combined in NADPH-dependent thioredoxin reductase C (NTRC), which contains an N-terminal thioredoxin reductase domain fused with a C-terminal thioredoxin domain. Rice NTRC exists in different oligomeric states, depending on the absence or presence of its NADPH cofactor. It has been suggested that the different oligomeric states may have diverse activity. Thus, the redox status of the chloroplast could influence the oligomeric state of NTRC and thereby its activity. We have characterized the oligomeric states of NTRC from barley (Hordeum vulgare L.). This also... (More)
Thioredoxin and thioredoxin reductase can regulate cell metabolism through redox regulation of disulfide bridges or through removal of H(2)O(2). These two enzymatic functions are combined in NADPH-dependent thioredoxin reductase C (NTRC), which contains an N-terminal thioredoxin reductase domain fused with a C-terminal thioredoxin domain. Rice NTRC exists in different oligomeric states, depending on the absence or presence of its NADPH cofactor. It has been suggested that the different oligomeric states may have diverse activity. Thus, the redox status of the chloroplast could influence the oligomeric state of NTRC and thereby its activity. We have characterized the oligomeric states of NTRC from barley (Hordeum vulgare L.). This also includes a structural model of the tetrameric NTRC derived from cryo-electron microscopy and single-particle reconstruction. We conclude that the tetrameric NTRC is a dimeric arrangement of two NTRC homodimers. Unlike that of rice NTRC, the quaternary structure of barley NTRC complexes is unaffected by addition of NADPH. The activity of NTRC was tested with two different enzyme assays. The N-terminal part of NTRC was tested in a thioredoxin reductase assay. A peroxide sensitive Mg-protoporphyrin IX monomethyl ester (MPE) cyclase enzyme system of the chlorophyll biosynthetic pathway was used to test the catalytic ability of both the N- and C-terminal parts of NTRC. The different oligomeric assembly states do not exhibit significantly different activities. Thus, it appears that the activities are independent of the oligomeric state of barley NTRC. (Less)
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organization
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type
Contribution to journal
publication status
published
subject
keywords
Protein Structure, Protein Conformation, Peroxides/chemistry, Oxidation-Reduction, NADP/*chemistry, Molecular Sequence Data, Molecular Conformation, Magnesium/chemistry, Hordeum/*enzymology, Dimerization, Cryoelectron Microscopy/methods, X-Ray/methods, Crystallography, Tertiary, Recombinant Proteins/chemistry, Thioredoxin-Disulfide Reductase/*chemistry, Thioredoxins/chemistry
in
Biochemistry
volume
50
issue
18
pages
3713 - 3723
publisher
The American Chemical Society
external identifiers
  • wos:000290056000014
  • scopus:79955589109
ISSN
0006-2960
DOI
10.1021/bi200058a
language
English
LU publication?
yes
id
30a562fa-600c-4752-9c8a-6c0fc9c11edb (old id 1988030)
date added to LUP
2015-09-30 13:24:19
date last changed
2017-08-27 03:02:48
@article{30a562fa-600c-4752-9c8a-6c0fc9c11edb,
  abstract     = {Thioredoxin and thioredoxin reductase can regulate cell metabolism through redox regulation of disulfide bridges or through removal of H(2)O(2). These two enzymatic functions are combined in NADPH-dependent thioredoxin reductase C (NTRC), which contains an N-terminal thioredoxin reductase domain fused with a C-terminal thioredoxin domain. Rice NTRC exists in different oligomeric states, depending on the absence or presence of its NADPH cofactor. It has been suggested that the different oligomeric states may have diverse activity. Thus, the redox status of the chloroplast could influence the oligomeric state of NTRC and thereby its activity. We have characterized the oligomeric states of NTRC from barley (Hordeum vulgare L.). This also includes a structural model of the tetrameric NTRC derived from cryo-electron microscopy and single-particle reconstruction. We conclude that the tetrameric NTRC is a dimeric arrangement of two NTRC homodimers. Unlike that of rice NTRC, the quaternary structure of barley NTRC complexes is unaffected by addition of NADPH. The activity of NTRC was tested with two different enzyme assays. The N-terminal part of NTRC was tested in a thioredoxin reductase assay. A peroxide sensitive Mg-protoporphyrin IX monomethyl ester (MPE) cyclase enzyme system of the chlorophyll biosynthetic pathway was used to test the catalytic ability of both the N- and C-terminal parts of NTRC. The different oligomeric assembly states do not exhibit significantly different activities. Thus, it appears that the activities are independent of the oligomeric state of barley NTRC.},
  author       = {Peterson Wulff, Ragna and Lundqvist, J. and Rutsdottir, Gudrun and Hansson, A. and Stenbaek, A. and Elmlund, D. and Elmlund, H. and Jensen, P. E. and Hansson, Mats},
  issn         = {0006-2960},
  keyword      = {Protein Structure,Protein Conformation,Peroxides/chemistry,Oxidation-Reduction,NADP/*chemistry,Molecular Sequence Data,Molecular Conformation,Magnesium/chemistry,Hordeum/*enzymology,Dimerization,Cryoelectron Microscopy/methods,X-Ray/methods,Crystallography,Tertiary,Recombinant Proteins/chemistry,Thioredoxin-Disulfide Reductase/*chemistry,Thioredoxins/chemistry},
  language     = {eng},
  number       = {18},
  pages        = {3713--3723},
  publisher    = {The American Chemical Society},
  series       = {Biochemistry},
  title        = {The activity of barley NADPH-dependent thioredoxin reductase C is independent of the oligomeric state of the protein: tetrameric structure determined by cryo-electron microscopy},
  url          = {http://dx.doi.org/10.1021/bi200058a},
  volume       = {50},
  year         = {2011},
}