NO Binding Induced Conformational Changes in a Truncated Hemoglobin from Mycobacterium tuberculosis

Masahiro Mukai, Yannick Ouellet, Hugues Ouellet, Michel Guertin, Syun-Ru Yeh

Research output: Contribution to journalArticle

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Abstract

The resonance Raman spectra of the NO-bound ferric derivatives of wild-type HbN and the B 10 Tyr → Phe mutant of HbN, a hemoglobin from Mycobacterium tuberculosis, were examined with both Soret and UV excitation. The Fe-N-O stretching and bending modes of the NO derivative of the wild-type protein were tentatively assigned at 591 and 579 cm-1, respectively. Upon B10 mutation, the Fe-NO stretching mode was slightly enhanced and the bending mode diminished in amplitude. In addition, the N-O stretching mode shifted from 1914 to 1908 cm-1. These data suggest that the B10 Tyr forms an H-bond(s) with the heme-bound NO and causes it to bend in the wild-type protein. To further investigate the interaction between the B10 Tyr and the heme-bound NO, we examined the UV Raman spectrum of the B10 Tyr by subtracting the B10 mutant spectrum from the wild-type spectrum. It was found that, upon NO binding to the ferric protein, the Y8a mode of the B10 Tyr shifted from 1616 to 1622 cm-1, confirming a direct interaction between the B 10 Tyr and the heme-bound NO. Furthermore, the Y8a mode of the other two Tyr residues at positions 16 and 72 that are remote from the heme was also affected by NO binding, suggesting that NO binding to the distal site of the heme triggers a large-scale conformational change that propagates through the pre-F helix loop to the E and B helices. This large-scale conformational change triggered by NO binding may play an important role in regulating the ligand binding properties and/or the chemical reactivity of HbN.

Original languageEnglish (US)
Pages (from-to)2764-2770
Number of pages7
JournalBiochemistry
Volume43
Issue number10
DOIs
StatePublished - Mar 16 2004

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Truncated Hemoglobins
Heme
Mycobacterium tuberculosis
Stretching
Raman scattering
Derivatives
Chemical reactivity
Proteins
Hemoglobins
Ligands
Mutation

ASJC Scopus subject areas

  • Biochemistry

Cite this

NO Binding Induced Conformational Changes in a Truncated Hemoglobin from Mycobacterium tuberculosis. / Mukai, Masahiro; Ouellet, Yannick; Ouellet, Hugues; Guertin, Michel; Yeh, Syun-Ru.

In: Biochemistry, Vol. 43, No. 10, 16.03.2004, p. 2764-2770.

Research output: Contribution to journalArticle

Mukai, Masahiro ; Ouellet, Yannick ; Ouellet, Hugues ; Guertin, Michel ; Yeh, Syun-Ru. / NO Binding Induced Conformational Changes in a Truncated Hemoglobin from Mycobacterium tuberculosis. In: Biochemistry. 2004 ; Vol. 43, No. 10. pp. 2764-2770.
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abstract = "The resonance Raman spectra of the NO-bound ferric derivatives of wild-type HbN and the B 10 Tyr → Phe mutant of HbN, a hemoglobin from Mycobacterium tuberculosis, were examined with both Soret and UV excitation. The Fe-N-O stretching and bending modes of the NO derivative of the wild-type protein were tentatively assigned at 591 and 579 cm-1, respectively. Upon B10 mutation, the Fe-NO stretching mode was slightly enhanced and the bending mode diminished in amplitude. In addition, the N-O stretching mode shifted from 1914 to 1908 cm-1. These data suggest that the B10 Tyr forms an H-bond(s) with the heme-bound NO and causes it to bend in the wild-type protein. To further investigate the interaction between the B10 Tyr and the heme-bound NO, we examined the UV Raman spectrum of the B10 Tyr by subtracting the B10 mutant spectrum from the wild-type spectrum. It was found that, upon NO binding to the ferric protein, the Y8a mode of the B10 Tyr shifted from 1616 to 1622 cm-1, confirming a direct interaction between the B 10 Tyr and the heme-bound NO. Furthermore, the Y8a mode of the other two Tyr residues at positions 16 and 72 that are remote from the heme was also affected by NO binding, suggesting that NO binding to the distal site of the heme triggers a large-scale conformational change that propagates through the pre-F helix loop to the E and B helices. This large-scale conformational change triggered by NO binding may play an important role in regulating the ligand binding properties and/or the chemical reactivity of HbN.",
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AB - The resonance Raman spectra of the NO-bound ferric derivatives of wild-type HbN and the B 10 Tyr → Phe mutant of HbN, a hemoglobin from Mycobacterium tuberculosis, were examined with both Soret and UV excitation. The Fe-N-O stretching and bending modes of the NO derivative of the wild-type protein were tentatively assigned at 591 and 579 cm-1, respectively. Upon B10 mutation, the Fe-NO stretching mode was slightly enhanced and the bending mode diminished in amplitude. In addition, the N-O stretching mode shifted from 1914 to 1908 cm-1. These data suggest that the B10 Tyr forms an H-bond(s) with the heme-bound NO and causes it to bend in the wild-type protein. To further investigate the interaction between the B10 Tyr and the heme-bound NO, we examined the UV Raman spectrum of the B10 Tyr by subtracting the B10 mutant spectrum from the wild-type spectrum. It was found that, upon NO binding to the ferric protein, the Y8a mode of the B10 Tyr shifted from 1616 to 1622 cm-1, confirming a direct interaction between the B 10 Tyr and the heme-bound NO. Furthermore, the Y8a mode of the other two Tyr residues at positions 16 and 72 that are remote from the heme was also affected by NO binding, suggesting that NO binding to the distal site of the heme triggers a large-scale conformational change that propagates through the pre-F helix loop to the E and B helices. This large-scale conformational change triggered by NO binding may play an important role in regulating the ligand binding properties and/or the chemical reactivity of HbN.

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