Publication
Title
Characterization of the heme pocket structure and ligand binding kinetics of nonsymbiotic hemoglobins from the model legume **Lotus japonicus**
Author
Abstract
Plant hemoglobins (Hbs) are found in nodules of legumes and actinorhizal plants but also in nonsymbiotic organs of monocots and dicots. Nonsymbiotic Hbs have been classified into two phylogenetic groups. Class 1 Hbs show an extremely high O2 affinity and are induced by hypoxia and nitric oxide (NO), whereas class 2 Hbs have moderate O2 affinity and are induced by cold and cytokinins. The functions of nonsymbiotic Hbs are still unclear, but some of them rely on the capacity of hemes to bind diatomic ligands and catalyze the NO dioxygenase reaction (oxyferrous Hb + NO → ferric Hb + nitrate). Moreover, NO may nitrosylate Cys residues of proteins. It is therefore important to determine the ligand binding properties of the hemes and the role of Cys residues. Here, we have addressed these issues with the two class 1 Hbs (LjGlb1-1 and LjGlb1-2) and the single class 2 Hb (LjGlb2) of Lotus japonicus, which is a model legume to facilitate the transfer of genetic and biochemical information into crops. We have used carbon monoxide (CO) as a model ligand and resonance Raman, laser flash photolysis, and stopped-flow spectroscopies to unveil major differences in the heme environments and ligand binding kinetics of the three proteins, which suggest non-redundant functions. In the deoxyferrous state, LjGlb1-1 is partially hexacoordinate, whereas LjGlb1-2 shows complete hexacoordination (behaving like class 2 Hbs) and LjGlb2 is mostly pentacoordinate (unlike other class 2 Hbs). LjGlb1-1 binds CO very strongly by stabilizing it through hydrogen bonding, but LjGlb1-2 and LjGlb2 show lower CO stabilization. The changes in CO stabilization would explain the different affinities of the three proteins for gaseous ligands. These affinities are determined by the dissociation rates and follow the order LjGlb1-1 > LjGlb1-2 > LjGlb2. Mutations LjGlb1-1 C78S and LjGlb1-2 C79S caused important alterations in protein dynamics and stability, indicating a structural role of those Cys residues, whereas mutation LjGlb1-1 C8S had a smaller effect. The three proteins and their mutant derivatives exhibited similarly high rates of NO consumption, which were due to NO dioxygenase activity of the hemes and not to nitrosylation of Cys residues.
Language
English
Source (journal)
Frontiers in plant science. - Place of publication unknown
Publication
Place of publication unknown : publisher unknown , 2017
ISSN
1664-462X
DOI
10.3389/FPLS.2017.00407
Volume/pages
8 (2017) , 14 p.
Article Reference
407
ISI
000398178400001
Pubmed ID
28421084
Medium
E-only publicatie
Full text (Publisher's DOI)
Full text (open access)
UAntwerpen
Faculty/Department
Research group
Project info
Towards new approaches in bioelectrochemistry – Targeted immobilization of globins on porous materials.
Publication type
Subject
Affiliation
Publications with a UAntwerp address
External links
Web of Science
Record
Identifier
Creation 09.03.2017
Last edited 04.03.2024
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