Resonance Raman spectroscopic study of nitrophorin 1, a nitric oxide-binding heme protein from Rhodnius prolixus, and its nitrosyl and cyano adducts

E. M. Maes, F. A. Walker, William "Bill" Montfort, R. S. Czernuszewicz

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Abstract

The resonance Raman (RR) spectra of nitrophorin 1 (NP1) from the saliva of the blood-sucking insect Rhodnius prolixus, in the absence and presence of nitric oxide (NO) and in the presence of cyanide (CN-), have been studied. The NP1 displayed RR spectra characteristic of six-coordinate high-spin (6cHS) ferric heme at room temperature and six-coordinate low-spin heme (6cLS) at low temperature (77 K). NO and CN- each bind to FeIII, both ligands forming 6cLS complexes with NP1. The FeIII-NO stretching and bending vibrational frequencies of nitrosyl NP1 were identified at 591 and 578 cm-1, respectively, on the basis of 15NO isotope shifts. These frequencies are typical of Fe-NO ferric heme proteins, indicating that the NP1 nitrosyl adduct has typical bond strength. Thus, the small NO release rate displayed by NP1 must be due to other protein interactions. Room and cryogenic temperature (77 K) RR spectroscopy and 13C, 15N, and 13C15N isotope substitutions have been used to determine vibrational mode frequencies associated with the FeIII-CN- bond for the cyano adducts at 454, 443, 397, and 357 cm-1. The results were analyzed by normal mode calculations to support the assignment of the modes and to assess the NO and CN- binding geometries. The observed isotope shifts for the cyano NP1 are smaller than expected and reveal vibrational coupling of FeIII-CN- modes with heme modes. We also find that the observed frequencies are consistent with the presence of a nearly linear FeIIICN- linkage (173°) coexisting with a population with a bent structure (155°).

Original languageEnglish (US)
Pages (from-to)11664-11672
Number of pages9
JournalJournal of the American Chemical Society
Volume123
Issue number47
DOIs
StatePublished - Nov 28 2001

Fingerprint

Rhodnius
Nitric oxide
Cyanides
Nitric Oxide
Isotopes
Heme
Raman scattering
Temperature
Hemeproteins
Vibrational spectra
Raman Spectrum Analysis
Cryogenics
Stretching
Raman spectroscopy
heme-binding protein
nitrophorin
Saliva
Blood
Substitution reactions
Insects

ASJC Scopus subject areas

  • Chemistry(all)

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Resonance Raman spectroscopic study of nitrophorin 1, a nitric oxide-binding heme protein from Rhodnius prolixus, and its nitrosyl and cyano adducts. / Maes, E. M.; Walker, F. A.; Montfort, William "Bill"; Czernuszewicz, R. S.

In: Journal of the American Chemical Society, Vol. 123, No. 47, 28.11.2001, p. 11664-11672.

Research output: Contribution to journalArticle

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abstract = "The resonance Raman (RR) spectra of nitrophorin 1 (NP1) from the saliva of the blood-sucking insect Rhodnius prolixus, in the absence and presence of nitric oxide (NO) and in the presence of cyanide (CN-), have been studied. The NP1 displayed RR spectra characteristic of six-coordinate high-spin (6cHS) ferric heme at room temperature and six-coordinate low-spin heme (6cLS) at low temperature (77 K). NO and CN- each bind to FeIII, both ligands forming 6cLS complexes with NP1. The FeIII-NO stretching and bending vibrational frequencies of nitrosyl NP1 were identified at 591 and 578 cm-1, respectively, on the basis of 15NO isotope shifts. These frequencies are typical of Fe-NO ferric heme proteins, indicating that the NP1 nitrosyl adduct has typical bond strength. Thus, the small NO release rate displayed by NP1 must be due to other protein interactions. Room and cryogenic temperature (77 K) RR spectroscopy and 13C, 15N, and 13C15N isotope substitutions have been used to determine vibrational mode frequencies associated with the FeIII-CN- bond for the cyano adducts at 454, 443, 397, and 357 cm-1. The results were analyzed by normal mode calculations to support the assignment of the modes and to assess the NO and CN- binding geometries. The observed isotope shifts for the cyano NP1 are smaller than expected and reveal vibrational coupling of FeIII-CN- modes with heme modes. We also find that the observed frequencies are consistent with the presence of a nearly linear FeIIICN- linkage (173°) coexisting with a population with a bent structure (155°).",
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N2 - The resonance Raman (RR) spectra of nitrophorin 1 (NP1) from the saliva of the blood-sucking insect Rhodnius prolixus, in the absence and presence of nitric oxide (NO) and in the presence of cyanide (CN-), have been studied. The NP1 displayed RR spectra characteristic of six-coordinate high-spin (6cHS) ferric heme at room temperature and six-coordinate low-spin heme (6cLS) at low temperature (77 K). NO and CN- each bind to FeIII, both ligands forming 6cLS complexes with NP1. The FeIII-NO stretching and bending vibrational frequencies of nitrosyl NP1 were identified at 591 and 578 cm-1, respectively, on the basis of 15NO isotope shifts. These frequencies are typical of Fe-NO ferric heme proteins, indicating that the NP1 nitrosyl adduct has typical bond strength. Thus, the small NO release rate displayed by NP1 must be due to other protein interactions. Room and cryogenic temperature (77 K) RR spectroscopy and 13C, 15N, and 13C15N isotope substitutions have been used to determine vibrational mode frequencies associated with the FeIII-CN- bond for the cyano adducts at 454, 443, 397, and 357 cm-1. The results were analyzed by normal mode calculations to support the assignment of the modes and to assess the NO and CN- binding geometries. The observed isotope shifts for the cyano NP1 are smaller than expected and reveal vibrational coupling of FeIII-CN- modes with heme modes. We also find that the observed frequencies are consistent with the presence of a nearly linear FeIIICN- linkage (173°) coexisting with a population with a bent structure (155°).

AB - The resonance Raman (RR) spectra of nitrophorin 1 (NP1) from the saliva of the blood-sucking insect Rhodnius prolixus, in the absence and presence of nitric oxide (NO) and in the presence of cyanide (CN-), have been studied. The NP1 displayed RR spectra characteristic of six-coordinate high-spin (6cHS) ferric heme at room temperature and six-coordinate low-spin heme (6cLS) at low temperature (77 K). NO and CN- each bind to FeIII, both ligands forming 6cLS complexes with NP1. The FeIII-NO stretching and bending vibrational frequencies of nitrosyl NP1 were identified at 591 and 578 cm-1, respectively, on the basis of 15NO isotope shifts. These frequencies are typical of Fe-NO ferric heme proteins, indicating that the NP1 nitrosyl adduct has typical bond strength. Thus, the small NO release rate displayed by NP1 must be due to other protein interactions. Room and cryogenic temperature (77 K) RR spectroscopy and 13C, 15N, and 13C15N isotope substitutions have been used to determine vibrational mode frequencies associated with the FeIII-CN- bond for the cyano adducts at 454, 443, 397, and 357 cm-1. The results were analyzed by normal mode calculations to support the assignment of the modes and to assess the NO and CN- binding geometries. The observed isotope shifts for the cyano NP1 are smaller than expected and reveal vibrational coupling of FeIII-CN- modes with heme modes. We also find that the observed frequencies are consistent with the presence of a nearly linear FeIIICN- linkage (173°) coexisting with a population with a bent structure (155°).

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