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Low-frequency mode activity of heme: femtosecond coherence spectroscopy of iron porphine halides and nitrophorin.

ABSTRACT: The low-frequency mode activity of metalloporphyrins has been studied for iron porphine-halides (Fe(P)(X), X = Cl, Br) and nitrophorin 4 (NP4) using femtosecond coherence spectroscopy (FCS) in combination with polarized resonance Raman spectroscopy and density functional theory (DFT). It is confirmed that the mode symmetry selection rules for FCS are the same as for Raman scattering and that both Franck-Condon and Jahn-Teller mode activities are observed for Fe(P)(X) under Soret resonance conditions. The DFT-calculated low-frequency (20-400 cm (-1)) modes, and their frequency shifts upon halide substitution, are in good agreement with experimental Raman and coherence data, so that mode assignments can be made. The doming mode is located at approximately 80 cm (-1) for Fe(P)(Cl) and at approximately 60 cm (-1) for Fe(P)(Br). NP4 is also studied with coherence techniques, and the NO-bound species of ferric and ferrous NP4 display a mode at approximately 30-40 cm (-1) that is associated with transient heme doming motion following NO photolysis. The coherence spectra of three ferric derivatives of NP4 with different degrees of heme ruffling distortion are also investigated. We find a mode at approximately 60 cm (-1) whose relative intensity in the coherence spectra depends quadratically on the magnitude of the ruffling distortion. To quantitatively account for this correlation, a new "distortion-induced" Raman enhancement mechanism is presented. This mechanism is unique to low-frequency "soft modes" of the molecular framework that can be distorted by environmental forces. These results demonstrate the potential of FCS as a sensitive probe of dynamic and functionally important nonplanar heme vibrational excitations that are induced by the protein environmental forces or by the chemical reactions in the aqueous phase.

SUBMITTER: Kubo M

PROVIDER: S-EPMC2765994 | biostudies-literature | 2008 Jul

REPOSITORIES: biostudies-literature

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Low-frequency mode activity of heme: femtosecond coherence spectroscopy of iron porphine halides and nitrophorin.

Kubo Minoru M Gruia Flaviu F Benabbas Abdelkrim A Barabanschikov Alexander A Montfort William R WR Maes Estelle M EM Champion Paul M PM

Journal of the American Chemical Society 20080703 30

The low-frequency mode activity of metalloporphyrins has been studied for iron porphine-halides (Fe(P)(X), X = Cl, Br) and nitrophorin 4 (NP4) using femtosecond coherence spectroscopy (FCS) in combination with polarized resonance Raman spectroscopy and density functional theory (DFT). It is confirmed that the mode symmetry selection rules for FCS are the same as for Raman scattering and that both Franck-Condon and Jahn-Teller mode activities are observed for Fe(P)(X) under Soret resonance condit ...[more]

PMID: 18597456

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Project description:Nitrophorin 3 (NP3) is the only one of the four major NO-binding heme proteins found in the saliva of the blood-sucking insect Rhodnius prolixus (also called the Kissing Bug) for which it has not been possible to obtain crystals of diffraction quality for structure determination by X-ray crystallography. Thus we have used NMR spectroscopy, mainly of the hyperfine-shifted ferriheme substituent resonances, to learn about the similarities and differences in the heme pocket and the iron active site of NP3 as compared to NP2, which has previously been well-characterized by both X-ray crystallography and NMR spectroscopy. Only one residue in the heme pocket differs between the two, F27 of NP2 is Y27 for NP3; in both cases this residue is expected to interact strongly with the 2-vinyl side chain of the B heme rotational isomer or the 4-vinyl of the A heme rotational isomer. Both the high-spin (S = 5/2) aquo complex, NP3-H(2)O, and the low-spin (S = 1/2) N-methylimidazole (NMeIm) complex of NP3 have been studied. It is found that the chemical shifts of the protons of both forms are similar to those of the corresponding NP2 complexes, but with minor differences that indicate a slightly different angle for the proximal histidine (H57) ligand plane. The B heme rotational isomer is preferred by both NP3 and NP2 in both spin states, but to a greater extent when phenylalanine is present at position 27 (A:B = 1:8 for NP2, 1:6 for NP3-Y27F, 1:4 for NP3, and 1:3 for NP2-F27Y). Careful analysis of the 5Me and 8Me shifts of the A and B isomers of the two high-spin nitrophorins leads to the conclusion that the heme environment for the two isomers differs in some way that cannot be explained at the present time. The kinetics of deprotonation of the high-spin complexes of NP2 and NP3 are very different, with NP2 giving well-resolved high-spin aquo and "low-spin" hydroxo proton NMR spectra until close to the end of the titration, while NP3 exhibits broadened (1)H NMR spectra indicative of an intermediate rate of exchange on the NMR timescale between the two forms throughout the titration. The heme methyl shifts of NP2-OH are similar in magnitude and spread to those of NP2-CN, while those of metmyoglobin-hydroxo complexes are much larger in magnitude but not spread. It is concluded that the hydroxo complex of NP2 is likely S = 1/2 with a mixed (d(XY))(2)(d(XZ),d(YZ))(3)/(d(xy))(1)(d(xz),d(yz))(4) electron configuration, while those of met-Mb-OH are likely S = 1/2,3/2 mixed spin systems.

Project description:Femtosecond vibrational coherence spectroscopy was used to investigate the low-frequency vibrational dynamics of the heme in the carbon monoxide oxidation activator protein (CooA) from the thermophilic anaerobic bacterium Carboxydothermus hydrogenoformans (Ch-CooA). Low frequency vibrational modes are important because they are excited by the ambient thermal bath (k(B)T = 200 cm(-1)) and participate in thermally activated barrier crossing events. However, such modes are nearly impossible to detect in the aqueous phase using traditional spectroscopic methods. Here, we present the low frequency coherence spectra of the ferric, ferrous, and CO-bound forms of Ch-CooA in order to compare the protein-induced heme distortions in its active and inactive states. Distortions take place predominantly along the coordinates of low-frequency modes because of their weak force constants, and such distortions are reflected in the intensity of the vibrational coherence signals. A strong mode near ~90 cm(-1) in the ferrous form of Ch-CooA is suggested to contain a large component of heme ruffling, consistent with the imidazole-bound ferrous heme crystal structure, which shows a significant protein-induced heme distortion along this coordinate. A mode observed at ~228 cm(-1) in the six-coordinate ferrous state is proposed to be the ?(Fe-His) stretching vibration. The observation of the Fe-His mode indicates that photolysis of the N-terminal ?-amino axial ligand takes place. This is followed by a rapid (~8.5 ps) transient absorption recovery, analogous to methionine rebinding in photolyzed ferrous cytochrome c. We have also studied CO photolysis in CooA, which revealed very strong photoproduct state coherent oscillations. The observation of heme-CO photoproduct oscillations is unusual because most other heme systems have CO rebinding kinetics that are too slow to make the measurement possible. The low frequency coherence spectrum of the CO-bound form of Ch-CooA shows a strong vibration at ~230 cm(-1) that is broadened and up-shifted compared to the ?(Fe-His) of Rr-CooA (216 cm(-1)). We propose that the stronger Fe-His bond is related to the enhanced thermal stability of Ch-CooA and that there is a smaller (time dependent) tilt of the histidine ring with respect to the heme plane in Ch-CooA. The appearance of strong modes at ~48 cm(-1) in both the ferrous and CO-bound forms of Ch-CooA is consistent with coupling of the heme doming distortion to the photolysis reaction in both samples. Upon CO binding and protein activation, a heme mode near 112 ± 5 cm(-1) disappears, probably indicating a decreased heme saddling distortion. This reflects changes in the heme environment and geometry that must be associated with the conformational transition activating the DNA-binding domain. Protein-specific DNA binding to the CO-bound form of Ch-CooA was also investigated, and although the CO rebinding kinetics are significantly perturbed, there are negligible changes in the low-frequency vibrational spectrum of the heme.

Dataset Information

Low-frequency mode activity of heme: femtosecond coherence spectroscopy of iron porphine halides and nitrophorin.

Publications

Low-frequency mode activity of heme: femtosecond coherence spectroscopy of iron porphine halides and nitrophorin.

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