5 results on '"Hegg EL"'
Search Results
2. Cytochrome c nitrite reductase from the bacterium Geobacter lovleyi represents a new NrfA subclass.
- Author
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Campeciño J, Lagishetty S, Wawrzak Z, Sosa Alfaro V, Lehnert N, Reguera G, Hu J, and Hegg EL
- Subjects
- Ammonium Compounds metabolism, Bacterial Proteins chemistry, Bacterial Proteins genetics, Crystallography, X-Ray, Cytochromes a1 chemistry, Cytochromes a1 genetics, Cytochromes c1 chemistry, Cytochromes c1 genetics, Geobacter chemistry, Geobacter genetics, Kinetics, Models, Molecular, Nitrate Reductases chemistry, Nitrate Reductases genetics, Nitrates metabolism, Phylogeny, Protein Conformation, Bacterial Proteins metabolism, Cytochromes a1 metabolism, Cytochromes c1 metabolism, Geobacter metabolism, Nitrate Reductases metabolism
- Abstract
Cytochrome c nitrite reductase (NrfA) catalyzes the reduction of nitrite to ammonium in the dissimilatory nitrate reduction to ammonium (DNRA) pathway, a process that competes with denitrification, conserves nitrogen, and minimizes nutrient loss in soils. The environmental bacterium Geobacter lovleyi has recently been recognized as a key driver of DNRA in nature, but its enzymatic pathway is still uncharacterized. To address this limitation, here we overexpressed, purified, and characterized G. lovleyi NrfA. We observed that the enzyme crystallizes as a dimer but remains monomeric in solution. Importantly, its crystal structure at 2.55-Å resolution revealed the presence of an arginine residue in the region otherwise occupied by calcium in canonical NrfA enzymes. The presence of EDTA did not affect the activity of G. lovleyi NrfA, and site-directed mutagenesis of this arginine reduced enzymatic activity to <3% of the WT levels. Phylogenetic analysis revealed four separate emergences of Arg-containing NrfA enzymes. Thus, the Ca
2+ -independent, Arg-containing NrfA from G. lovleyi represents a new subclass of cytochrome c nitrite reductase. Most genera from the exclusive clades of Arg-containing NrfA proteins are also represented in clades containing Ca2+ -dependent enzymes, suggesting convergent evolution., Competing Interests: Conflict of interest—The authors declare no conflicts of interest in regards to this work., (© 2020 Campeciño et al.)- Published
- 2020
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3. Cox15 interacts with the cytochrome bc 1 dimer within respiratory supercomplexes as well as in the absence of cytochrome c oxidase.
- Author
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Herwaldt EJ, Rivett ED, White AJ, and Hegg EL
- Subjects
- Cytochrome-c Oxidase Deficiency, Heme analogs & derivatives, Saccharomyces cerevisiae, Electron Transport Complex III metabolism, Electron Transport Complex IV metabolism, Membrane Proteins metabolism, Saccharomyces cerevisiae Proteins metabolism
- Abstract
The heme a molecule is an obligatory cofactor in the terminal enzyme complex of the electron transport chain, cytochrome c oxidase. Heme a is synthesized from heme o by a multi-spanning inner membrane protein, heme a synthase (Cox15 in the yeast Saccharomyces cerevisiae ). The insertion of heme a is critical for cytochrome c oxidase function and assembly, but this process has not been fully elucidated. To improve our understanding of heme a insertion into cytochrome c oxidase, here we investigated the protein-protein interactions that involve Cox15 in S. cerevisiae In addition to observing Cox15 in homooligomeric complexes, we found that a portion of Cox15 also associates with the mitochondrial respiratory supercomplexes. When supercomplex formation was abolished, as in the case of stalled cytochrome bc
1 or cytochrome c oxidase assembly, Cox15 maintained an interaction with select proteins from both respiratory complexes. In the case of stalled cytochrome bc1 assembly, Cox15 interacted with the late-assembling cytochrome c oxidase subunit, Cox13. When cytochrome c oxidase assembly was stalled, Cox15 unexpectedly maintained its interaction with the cytochrome bc1 protein, Cor1. Our results indicate that Cox15 and Cor1 continue to interact in the cytochrome bc1 dimer even in the absence of supercomplexes or when the supercomplexes are destabilized. These findings reveal that Cox15 not only associates with respiratory supercomplexes, but also interacts with the cytochrome bc1 dimer even in the absence of cytochrome c oxidase., (© 2018 Herwaldt et al.)- Published
- 2018
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4. Mechanism of proton transfer in [FeFe]-hydrogenase from Clostridium pasteurianum.
- Author
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Cornish AJ, Gärtner K, Yang H, Peters JW, and Hegg EL
- Subjects
- Amino Acid Sequence, Biological Transport, Catalysis, Glutamic Acid chemistry, Iron-Sulfur Proteins chemistry, Kinetics, Molecular Conformation, Molecular Sequence Data, Mutagenesis, Site-Directed, Protons, Sequence Homology, Amino Acid, Zinc chemistry, Clostridium enzymology, Hydrogenase chemistry, Iron chemistry
- Abstract
[FeFe]-Hydrogenases are complex metalloproteins that catalyze the reversible reduction of protons to molecular hydrogen utilizing a unique diiron subcluster bridged to a [4Fe4S] subcluster. Extensive studies have concentrated on the nature and catalytic activity of the active site, yet relatively little information is available concerning the mechanism of proton transport that is required for this activity. Previously, structural characterization of [FeFe]-hydrogenase from Clostridium pasteurianum indicated a potential proton transport pathway involving four residues (Cys-299, Glu-279, Ser-319, and Glu-282) that connect the active site to the enzyme surface. Here, we demonstrate that substitution of any of these residues resulted in a drastic reduction in hydrogenase activity relative to the native enzyme, supporting the importance of these residues in catalysis. Inhibition studies of native and amino acid-substituted enzymes revealed that Zn(2+) specifically blocked proton transfer by binding to Glu-282, confirming the role of this residue in the identified pathway. In addition, all four of these residues are strictly conserved, suggesting that they may form a proton transport pathway that is common to all [FeFe]-hydrogenases.
- Published
- 2011
- Full Text
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5. Regulation of the heme A biosynthetic pathway: differential regulation of heme A synthase and heme O synthase in Saccharomyces cerevisiae.
- Author
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Wang Z, Wang Y, and Hegg EL
- Subjects
- Alkyl and Aryl Transferases genetics, DNA-Binding Proteins, Electron Transport Complex IV genetics, Electron Transport Complex IV metabolism, Gene Expression Regulation, Fungal, Heme biosynthesis, Isoenzymes genetics, Isoenzymes metabolism, Membrane Proteins genetics, Promoter Regions, Genetic genetics, RNA, Messenger genetics, Saccharomyces cerevisiae genetics, Saccharomyces cerevisiae Proteins genetics, Trans-Activators genetics, Trans-Activators metabolism, Transcription Factors, Alkyl and Aryl Transferases metabolism, Gene Expression Regulation, Enzymologic genetics, Heme analogs & derivatives, Membrane Proteins metabolism, Saccharomyces cerevisiae enzymology, Saccharomyces cerevisiae Proteins metabolism
- Abstract
The assembly and activity of cytochrome c oxidase is dependent on the availability of heme A, one of its essential cofactors. In eukaryotes, two inner mitochondrial membrane proteins, heme O synthase (Cox10) and heme A synthase (Cox15), are required for heme A biosynthesis. In this report, we demonstrate that in Saccharomyces cerevisiae the transcription of COX15 is regulated by Hap1, a transcription factor whose activity is positively controlled by intracellular heme concentration. Conversely, COX10, the physiological partner of COX15, does not share the same regulatory mechanism with COX15. Interestingly, protein quantification identified an 8:1 protein ratio between Cox15 and Cox10. Together, these results suggest that heme A synthase and/or heme O synthase might play a new, unidentified role in addition to heme A biosynthesis.
- Published
- 2009
- Full Text
- View/download PDF
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