Carbon-13-NMR of peptides and proteins

We have synthesized and studied via solid-state NMR, Mossbauer spectroscopy, single-crystal X-ray diffraction, and density functional theory the following Fe−O2 analogue metalloporphyrins:  Fe(5,10,15,20-tetraphenylporphyrinate) (nitrosobenzene)(1-methylimidazole); Fe(5,10,15,20-tetraphenylporphyrinate) (nitrosobenzene)(pyridine); Fe(5,10,15,20-tetraphenylporphyrinate)(4-nitroso-N,N-dimethylaniline)(pyridine); Fe(2,3,7,8,12,13,17,18-octaethylporphyrinate) (nitrosobenzene)(1-methylimidazole) and Co(2,3,7,8,12,13,17,18-octaethylporphyrinate)(NO). Our results show that the porphyrin rings of the two tetraphenylporphyrins containing pyridine are ruffled while the other three compounds are planar:  reasons for this are discussed. The solid-state NMR and Mossbauer spectroscopic results are well reproduced by the DFT calculations, which then enable the testing of various models of Fe−O2 bonding in metalloporphyrins and metalloproteins. We find no evidence for two binding sites in oxypicket fence porphyrin, chara...

Solid-State NMR, Crystallographic and Density Functional Theory Investigation of Fe−CO and Fe−CO Analogue Metalloporphyrins and Metalloproteins†

Publisher Summary This chapter discusses the observation of numerous single-carbon resonances in natural-abundance, 13C nuclear magnetic resonance (NMR) spectra of small native proteins in solution, and with techniques and procedures that facilitate the use of the observed resonances for probing the environments of individual amino acid residues of native proteins in solution. The coverage is restricted mainly to the use of nonprotonated aromatic carbon resonances. All spectra presented in the chapter are proton decoupled natural-abundance 13C NMR spectra obtained at 15.18 MHz (14.2 kG), on a spectrometer equipped with a 20-mm probe. The chapter is concerned with three parameters of 13C relaxation: spin–lattice relaxation, spin–spin relaxation, and the nuclear Overhauser effect (NOE). The chapter also defines the NOE as the ratio of the integrated intensity of a 13C resonance under conditions of proton decoupling and the integrated intensity of the same resonance when no proton decoupling is applied. It also defines the NOE of a 13C resonance as the ratio of intensities with and without proton decoupling.

[19] Carbon-13 nuclear magnetic resonance: New techniques

Publisher Summary This chapter discusses the history of nuclear magnetic resonance (NMR) of amino acids, peptides, and proteins. Biological NMR technique is a very powerful one and has demonstrated its ability to elucidate the structures of smaller molecules to the extent that its use is essential for most organic chemists, but when applied to biological macromolecules it becomes subject to three major disadvantages that is described in the chapter. This chapter illustrates that overcoming the problems of resolution and assignment is thus a necessary feature of any NMR approach to macromolecules and it is true to say that many papers that reach the literature are reports of work that never really succeeded in overcoming both. This chapter briefly discusses the chief methods used in overcoming the resolution problem. This problem is most marked in proton spectroscopy; the much larger chemical shift range found, for example, in C spectra shifts the main problem from resolution to assignment and signal-to-noise ratio.

NMR of Amino Acids, Peptides, and Proteins (1977–1979)

Catalysis of para hydroxylation of aniline was measured for human ferrihemoglobin and various derivatives in a reconstituted system consisting of the appropriate hemoprotein (at 4 microM heme), reduced nicotinamide adenine dinucleotide phosphate (NADPH), cytochrome P-450 reductase, and aniline under atmospheric O2. The isolated subunits of hemoglobin (alpha 3+ and beta 3+4) were prepared by treatment with p-(hydroxymercuri)benzoate. Semihemoglobin (alpha heme2 beta 02) was prepared from ferrihemoglobin and apohemoglobin. Converse valency hybrids alpha 3+2(beta 2+-CO)2 and (alpha 2+-CO)2 beta 3+2 were prepared from appropriately ligated alpha and beta subunits. After chromatography, the hemoglobin derivatives were characterized by visible and 1H NMR spectroscopy and electrophoresis. At the same concentration of aniline, the alpha and beta subunits were much less active than the normal tetramer. alpha-Semihemoglobin and the alpha 3+2(beta 2+-CO)2 hybrid also displayed lower hydroxylase activity. The (alpha 2+-CO)2 beta 3+2 hybrid was about as active as normal alpha 3+2 beta 3+2. This result suggests that the activity of tetrameric hemoglobin primarily involves the beta subunits. Also transfer of the beta subunits from the beta 4 molecular environment to the alpha 2 beta 2 state enhances their monooxygenase activity approximately 15-fold. The hemoglobin derivatives were differently susceptible to substrate inhibition, the beta 4 species being most sensitive. Estimates of Vmax from the linear portions of the corresponding Lineweaver-Burk plots showed agreement within a factor of 2.5 for all of the hemoglobin derivatives, suggesting that the intrinsic O2-activating capacities of the derivatives are similar.(ABSTRACT TRUNCATED AT 250 WORDS)

Monooxygenase activity of human hemoglobin: role of quaternary structure in the preponderant activity of the .beta. subunits within the tetramer

Carbon-13 NMR studies of C2H5N13C binding to various hemoproteins.

Carbon-13 NMR studies of C2H5N13C bound to hemoproteins — Evidence for a different distal proteic environment in β-chains either isolated or within human hemoglobin

B. Cendrier

Papers

Carbon-13 NMR studies of C2H5N13C binding to various hemoproteins.

Carbon-13 NMR studies of C2H5N13C bound to hemoproteins — Evidence for a different distal proteic environment in β-chains either isolated or within human hemoglobin