Heme oxidation in a chimeric protein of the α-selective Neisseriae meningitidis heme oxygenase with the distal helix of the δ-selective Pseudomonas aeruginosa

Rahul Deshmukh, Yuhong Zeng, Lena M. Furci, Hong Wei Huang, Bailey N. Morgan, Suzanne Sander, Aileen Y. Alontaga, Richard A. Bunce, Pierre Moenne-Loccoz, Mario Rivera, Angela Wilks

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Abstract

Heme oxygenases from the bacterial pathogens Neisseriae meningitidis (nm-HO) and Pseudomonas aeruginosa (pa-HO) share significant sequence identity (37%). In nm-HO, biliverdin IXα is the sole product of the reaction, whereas pa-HO yields predominantly biliverdin IXδ. We have previously shown by NMR that the in-plane conformation of the heme in pa-HO is significantly different from that of nm-HO as a result of distinct interactions of the heme propionates with the protein scaffold [Caignan, G. A., Deshmukh, R., Wilks, A., Zeng, Y., Huang, H. W., Moenne-Loccoz, P., Bunce, R. A., Eastman, M. A., and Rivera, M. (2002) J. Am. Chem. Soc. 124, 14879-14892]. In the report presented here, we have extended these studies to investigate the role of the distal helix by preparing a chimera of nm-HO (nm-HOch), in which distal helix residues 107-142 of nm-HO have been replaced with the corresponding residues of the δ-regioselective pa-HO (112-147). Electronic absorption spectra, resonance Raman and FTIR spectroscopic studies confirm that the orientation and hydrogen bonding properties of the proximal His ligand are not significantly altered in the chimera relative those of the wild-type proteins. The catalytic turnover of the nm-HOch-heme complex yields almost exclusively α-biliverdin and a small but reproducible amount of δ-biliverdin. NMR spectroscopic studies reveal that the altered regioselectivity in the chimeric protein likely stems from a dynamic equilibrium between two alternate in-plane conformations of the heme (in-plane heme disorder). Replacement of K16 with Ala and Met31 with Lys in the chimeric protein in an effort to tune key polypeptide-heme propionate contacts largely stabilizes the in-plane conformer conducive to δ-meso hydroxylation.

Original languageEnglish (US)
Pages (from-to)13713-13723
Number of pages11
JournalBiochemistry
Volume44
Issue number42
DOIs
StatePublished - Oct 25 2005

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Heme Oxygenase (Decyclizing)
Neisseria meningitidis
Heme
Biliverdine
Pseudomonas aeruginosa
Oxidation
Proteins
Propionates
Conformations
Nuclear magnetic resonance
Regioselectivity
Hydroxylation
Pathogens
Fourier Transform Infrared Spectroscopy
Hydrogen Bonding
Scaffolds
Absorption spectra
Hydrogen bonds
Ligands
Peptides

ASJC Scopus subject areas

  • Biochemistry

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Heme oxidation in a chimeric protein of the α-selective Neisseriae meningitidis heme oxygenase with the distal helix of the δ-selective Pseudomonas aeruginosa. / Deshmukh, Rahul; Zeng, Yuhong; Furci, Lena M.; Huang, Hong Wei; Morgan, Bailey N.; Sander, Suzanne; Alontaga, Aileen Y.; Bunce, Richard A.; Moenne-Loccoz, Pierre; Rivera, Mario; Wilks, Angela.

In: Biochemistry, Vol. 44, No. 42, 25.10.2005, p. 13713-13723.

Research output: Contribution to journalArticle

Deshmukh, R, Zeng, Y, Furci, LM, Huang, HW, Morgan, BN, Sander, S, Alontaga, AY, Bunce, RA, Moenne-Loccoz, P, Rivera, M & Wilks, A 2005, 'Heme oxidation in a chimeric protein of the α-selective Neisseriae meningitidis heme oxygenase with the distal helix of the δ-selective Pseudomonas aeruginosa', Biochemistry, vol. 44, no. 42, pp. 13713-13723. https://doi.org/10.1021/bi050810t
Deshmukh, Rahul ; Zeng, Yuhong ; Furci, Lena M. ; Huang, Hong Wei ; Morgan, Bailey N. ; Sander, Suzanne ; Alontaga, Aileen Y. ; Bunce, Richard A. ; Moenne-Loccoz, Pierre ; Rivera, Mario ; Wilks, Angela. / Heme oxidation in a chimeric protein of the α-selective Neisseriae meningitidis heme oxygenase with the distal helix of the δ-selective Pseudomonas aeruginosa. In: Biochemistry. 2005 ; Vol. 44, No. 42. pp. 13713-13723.
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abstract = "Heme oxygenases from the bacterial pathogens Neisseriae meningitidis (nm-HO) and Pseudomonas aeruginosa (pa-HO) share significant sequence identity (37{\%}). In nm-HO, biliverdin IXα is the sole product of the reaction, whereas pa-HO yields predominantly biliverdin IXδ. We have previously shown by NMR that the in-plane conformation of the heme in pa-HO is significantly different from that of nm-HO as a result of distinct interactions of the heme propionates with the protein scaffold [Caignan, G. A., Deshmukh, R., Wilks, A., Zeng, Y., Huang, H. W., Moenne-Loccoz, P., Bunce, R. A., Eastman, M. A., and Rivera, M. (2002) J. Am. Chem. Soc. 124, 14879-14892]. In the report presented here, we have extended these studies to investigate the role of the distal helix by preparing a chimera of nm-HO (nm-HOch), in which distal helix residues 107-142 of nm-HO have been replaced with the corresponding residues of the δ-regioselective pa-HO (112-147). Electronic absorption spectra, resonance Raman and FTIR spectroscopic studies confirm that the orientation and hydrogen bonding properties of the proximal His ligand are not significantly altered in the chimera relative those of the wild-type proteins. The catalytic turnover of the nm-HOch-heme complex yields almost exclusively α-biliverdin and a small but reproducible amount of δ-biliverdin. NMR spectroscopic studies reveal that the altered regioselectivity in the chimeric protein likely stems from a dynamic equilibrium between two alternate in-plane conformations of the heme (in-plane heme disorder). Replacement of K16 with Ala and Met31 with Lys in the chimeric protein in an effort to tune key polypeptide-heme propionate contacts largely stabilizes the in-plane conformer conducive to δ-meso hydroxylation.",
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T1 - Heme oxidation in a chimeric protein of the α-selective Neisseriae meningitidis heme oxygenase with the distal helix of the δ-selective Pseudomonas aeruginosa

AU - Deshmukh, Rahul

AU - Zeng, Yuhong

AU - Furci, Lena M.

AU - Huang, Hong Wei

AU - Morgan, Bailey N.

AU - Sander, Suzanne

AU - Alontaga, Aileen Y.

AU - Bunce, Richard A.

AU - Moenne-Loccoz, Pierre

AU - Rivera, Mario

AU - Wilks, Angela

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N2 - Heme oxygenases from the bacterial pathogens Neisseriae meningitidis (nm-HO) and Pseudomonas aeruginosa (pa-HO) share significant sequence identity (37%). In nm-HO, biliverdin IXα is the sole product of the reaction, whereas pa-HO yields predominantly biliverdin IXδ. We have previously shown by NMR that the in-plane conformation of the heme in pa-HO is significantly different from that of nm-HO as a result of distinct interactions of the heme propionates with the protein scaffold [Caignan, G. A., Deshmukh, R., Wilks, A., Zeng, Y., Huang, H. W., Moenne-Loccoz, P., Bunce, R. A., Eastman, M. A., and Rivera, M. (2002) J. Am. Chem. Soc. 124, 14879-14892]. In the report presented here, we have extended these studies to investigate the role of the distal helix by preparing a chimera of nm-HO (nm-HOch), in which distal helix residues 107-142 of nm-HO have been replaced with the corresponding residues of the δ-regioselective pa-HO (112-147). Electronic absorption spectra, resonance Raman and FTIR spectroscopic studies confirm that the orientation and hydrogen bonding properties of the proximal His ligand are not significantly altered in the chimera relative those of the wild-type proteins. The catalytic turnover of the nm-HOch-heme complex yields almost exclusively α-biliverdin and a small but reproducible amount of δ-biliverdin. NMR spectroscopic studies reveal that the altered regioselectivity in the chimeric protein likely stems from a dynamic equilibrium between two alternate in-plane conformations of the heme (in-plane heme disorder). Replacement of K16 with Ala and Met31 with Lys in the chimeric protein in an effort to tune key polypeptide-heme propionate contacts largely stabilizes the in-plane conformer conducive to δ-meso hydroxylation.

AB - Heme oxygenases from the bacterial pathogens Neisseriae meningitidis (nm-HO) and Pseudomonas aeruginosa (pa-HO) share significant sequence identity (37%). In nm-HO, biliverdin IXα is the sole product of the reaction, whereas pa-HO yields predominantly biliverdin IXδ. We have previously shown by NMR that the in-plane conformation of the heme in pa-HO is significantly different from that of nm-HO as a result of distinct interactions of the heme propionates with the protein scaffold [Caignan, G. A., Deshmukh, R., Wilks, A., Zeng, Y., Huang, H. W., Moenne-Loccoz, P., Bunce, R. A., Eastman, M. A., and Rivera, M. (2002) J. Am. Chem. Soc. 124, 14879-14892]. In the report presented here, we have extended these studies to investigate the role of the distal helix by preparing a chimera of nm-HO (nm-HOch), in which distal helix residues 107-142 of nm-HO have been replaced with the corresponding residues of the δ-regioselective pa-HO (112-147). Electronic absorption spectra, resonance Raman and FTIR spectroscopic studies confirm that the orientation and hydrogen bonding properties of the proximal His ligand are not significantly altered in the chimera relative those of the wild-type proteins. The catalytic turnover of the nm-HOch-heme complex yields almost exclusively α-biliverdin and a small but reproducible amount of δ-biliverdin. NMR spectroscopic studies reveal that the altered regioselectivity in the chimeric protein likely stems from a dynamic equilibrium between two alternate in-plane conformations of the heme (in-plane heme disorder). Replacement of K16 with Ala and Met31 with Lys in the chimeric protein in an effort to tune key polypeptide-heme propionate contacts largely stabilizes the in-plane conformer conducive to δ-meso hydroxylation.

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