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Conformational Dynamics of Phytoglobin BvPgb1.2 from Beta vulgaris ssp. vulgaris

Christensen, Simon LU orcid ; Stenström, Olof LU ; Akke, Mikael LU orcid and Bülow, Leif LU (2023) In International Journal of Molecular Sciences 24(4).
Abstract

Plant hemoglobins, often referred to as phytoglobins, play important roles in abiotic stress tolerance. Several essential small physiological metabolites can be bound to these heme proteins. In addition, phytoglobins can catalyze a range of different oxidative reactions in vivo. These proteins are often oligomeric, but the degree and relevance of subunit interactions are largely unknown. In this study, we delineate which residues are involved in dimer formation of a sugar beet phytoglobin type 1.2 (BvPgb1.2) using NMR relaxation experiments. E. coli cells harboring a phytoglobin expression vector were cultivated in isotope-labeled (2H, 13C and 15N) M9 medium. The triple-labeled protein was purified to... (More)

Plant hemoglobins, often referred to as phytoglobins, play important roles in abiotic stress tolerance. Several essential small physiological metabolites can be bound to these heme proteins. In addition, phytoglobins can catalyze a range of different oxidative reactions in vivo. These proteins are often oligomeric, but the degree and relevance of subunit interactions are largely unknown. In this study, we delineate which residues are involved in dimer formation of a sugar beet phytoglobin type 1.2 (BvPgb1.2) using NMR relaxation experiments. E. coli cells harboring a phytoglobin expression vector were cultivated in isotope-labeled (2H, 13C and 15N) M9 medium. The triple-labeled protein was purified to homogeneity using two chromatographic steps. Two forms of BvPgb1.2 were examined, the oxy-form and the more stable cyanide-form. Using three-dimensional triple-resonance NMR experiments, sequence-specific assignments for CN-bound BvPgb1.2 were achieved for 137 backbone amide cross-peaks in the 1H-15N TROSY spectrum, which amounts to 83% of the total number of 165 expected cross-peaks. A large proportion of the non-assigned residues are located in α-helixes G and H, which are proposed to be involved in protein dimerization. Such knowledge around dimer formation will be instrumental for developing a better understanding of phytoglobins’ roles in planta.

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author
; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
amino acid labeling, hemoglobin, NMR relaxation, phytoglobin, subunit interactions, sugar beet
in
International Journal of Molecular Sciences
volume
24
issue
4
article number
3973
publisher
MDPI AG
external identifiers
  • pmid:36835381
  • scopus:85149298258
ISSN
1661-6596
DOI
10.3390/ijms24043973
language
English
LU publication?
yes
id
29f7ef9a-d479-47e0-b732-1913c06eaa67
date added to LUP
2024-01-12 14:21:36
date last changed
2024-04-13 07:49:05
@article{29f7ef9a-d479-47e0-b732-1913c06eaa67,
  abstract     = {{<p>Plant hemoglobins, often referred to as phytoglobins, play important roles in abiotic stress tolerance. Several essential small physiological metabolites can be bound to these heme proteins. In addition, phytoglobins can catalyze a range of different oxidative reactions in vivo. These proteins are often oligomeric, but the degree and relevance of subunit interactions are largely unknown. In this study, we delineate which residues are involved in dimer formation of a sugar beet phytoglobin type 1.2 (BvPgb1.2) using NMR relaxation experiments. E. coli cells harboring a phytoglobin expression vector were cultivated in isotope-labeled (<sup>2</sup>H, <sup>13</sup>C and <sup>15</sup>N) M9 medium. The triple-labeled protein was purified to homogeneity using two chromatographic steps. Two forms of BvPgb1.2 were examined, the oxy-form and the more stable cyanide-form. Using three-dimensional triple-resonance NMR experiments, sequence-specific assignments for CN-bound BvPgb1.2 were achieved for 137 backbone amide cross-peaks in the <sup>1</sup>H-<sup>15</sup>N TROSY spectrum, which amounts to 83% of the total number of 165 expected cross-peaks. A large proportion of the non-assigned residues are located in α-helixes G and H, which are proposed to be involved in protein dimerization. Such knowledge around dimer formation will be instrumental for developing a better understanding of phytoglobins’ roles in planta.</p>}},
  author       = {{Christensen, Simon and Stenström, Olof and Akke, Mikael and Bülow, Leif}},
  issn         = {{1661-6596}},
  keywords     = {{amino acid labeling; hemoglobin; NMR relaxation; phytoglobin; subunit interactions; sugar beet}},
  language     = {{eng}},
  number       = {{4}},
  publisher    = {{MDPI AG}},
  series       = {{International Journal of Molecular Sciences}},
  title        = {{Conformational Dynamics of Phytoglobin BvPgb1.2 from Beta vulgaris ssp. vulgaris}},
  url          = {{http://dx.doi.org/10.3390/ijms24043973}},
  doi          = {{10.3390/ijms24043973}},
  volume       = {{24}},
  year         = {{2023}},
}