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1H and 13C NMR spectroscopy of paramagnetic low-spin model heme complexes.
STItUCTURE/FUNCTION
145
B090
lH AND 13C NMR SPECTROSCOPY OF PARAMAGNETIC LOW-SPIN MODEL HEME COMPLEXES. U. Simonis’, H. Tan’, F. A. Walkerb, a Department,of Chemistryand Biochemistry,San Francisco State University,San Francisco, CA 94132. b Deparnnent of Chemistry, Universityof Arizona, Tucson, AZ 85721. ‘H and 13C NMR spectroscopy, using hyperfine shifts and paramagnetic relaxation, provide valuable information on the electronic, magnetic, and molecular structure properties of the active sites of paramagnetic proteins, such as heme proteins, and their corresponding model complexes’. Access to this information was hampered for some time due to the assumption that modern one- and twodimensional NMR experiments were not suitable for the resonance and structure assignment of paramagnetic molecules. It was believed that the short relaxation times of the complexes would render cross peak intensity undetectable. However, we showed recently that two-dimensional ‘H COSY and NOESY NMR experiments can be used for the unambiguous assignment of the pyrrole proton resonances of N-methylimidazole complexes of mono-ortho substituted tetraphenylporphyrinatoiron(II1). This assignment has the potential to clearly delineate the pattern of spin-delocalization in both natural and synthetic hemes and to describe the electron density distribution in the porphyrin ?r-orbitais. The COSY and NOESY correlations observed are consistent with a newly developed model which predicts the electron density distribution for the porphyrin a-orbitals in these complexes. To investigate if such a model can also be obtained for the assignment of the pyrrole proton resonances in mono-para or mono-meta substituted tetraphenylporphyrinatoiron(II1) complexes, we have synthesized several complexes bearing a unique substituent in para or one meta position of one phenyl ring. IH COSY and NOESY spectra of these complexes indeed reveal cross peaks consistent with those predicted for the electron density distribution in the porphyrin n-orbitais, modified by the nature of the substituent. 13C NMR spectroscopy is also a very sensitive and accurate tool for mappin the spin density distribution at the porphyrin carbon atoms. A complete ps C resonance assignment, which is a crucial requirement for the investigation of the nature of r-spin distribution at the p-pyrrole positions, can be achieved by a combination of one-dimensional polarization transfer (DEPT) and two-dimensional ‘H-13C correlated spectroscopy (HETCOR). The correlation pattern observed in both the proton and carbon 2D maps, as well as the experimental considerations necessary for obtaining heteronuclear cross correIations among hyperfine-shifted resonances of the paramagnetic complexes will be discussed in detail. 1. I. Bert& C. Luchinat, L. Messori, and M. Vase&J. Am. Chem. SW., 111,792O (1989). 2. Q. Lin, U. Simonis, A. R. Tipton, C. R. NotvQl, and F. A. Walker, Inorg. Chem., 31, 4216 (1992). 3. U. Simonis, F. A. Walker, and Q. Lin, J. Cdl. Biochm., 17C, 301 (1993).
145
B090
lH AND 13C NMR SPECTROSCOPY OF PARAMAGNETIC LOW-SPIN MODEL HEME COMPLEXES. U. Simonis’, H. Tan’, F. A. Walkerb, a Department,of Chemistryand Biochemistry,San Francisco State University,San Francisco, CA 94132. b Deparnnent of Chemistry, Universityof Arizona, Tucson, AZ 85721. ‘H and 13C NMR spectroscopy, using hyperfine shifts and paramagnetic relaxation, provide valuable information on the electronic, magnetic, and molecular structure properties of the active sites of paramagnetic proteins, such as heme proteins, and their corresponding model complexes’. Access to this information was hampered for some time due to the assumption that modern one- and twodimensional NMR experiments were not suitable for the resonance and structure assignment of paramagnetic molecules. It was believed that the short relaxation times of the complexes would render cross peak intensity undetectable. However, we showed recently that two-dimensional ‘H COSY and NOESY NMR experiments can be used for the unambiguous assignment of the pyrrole proton resonances of N-methylimidazole complexes of mono-ortho substituted tetraphenylporphyrinatoiron(II1). This assignment has the potential to clearly delineate the pattern of spin-delocalization in both natural and synthetic hemes and to describe the electron density distribution in the porphyrin ?r-orbitais. The COSY and NOESY correlations observed are consistent with a newly developed model which predicts the electron density distribution for the porphyrin a-orbitals in these complexes. To investigate if such a model can also be obtained for the assignment of the pyrrole proton resonances in mono-para or mono-meta substituted tetraphenylporphyrinatoiron(II1) complexes, we have synthesized several complexes bearing a unique substituent in para or one meta position of one phenyl ring. IH COSY and NOESY spectra of these complexes indeed reveal cross peaks consistent with those predicted for the electron density distribution in the porphyrin n-orbitais, modified by the nature of the substituent. 13C NMR spectroscopy is also a very sensitive and accurate tool for mappin the spin density distribution at the porphyrin carbon atoms. A complete ps C resonance assignment, which is a crucial requirement for the investigation of the nature of r-spin distribution at the p-pyrrole positions, can be achieved by a combination of one-dimensional polarization transfer (DEPT) and two-dimensional ‘H-13C correlated spectroscopy (HETCOR). The correlation pattern observed in both the proton and carbon 2D maps, as well as the experimental considerations necessary for obtaining heteronuclear cross correIations among hyperfine-shifted resonances of the paramagnetic complexes will be discussed in detail. 1. I. Bert& C. Luchinat, L. Messori, and M. Vase&J. Am. Chem. SW., 111,792O (1989). 2. Q. Lin, U. Simonis, A. R. Tipton, C. R. NotvQl, and F. A. Walker, Inorg. Chem., 31, 4216 (1992). 3. U. Simonis, F. A. Walker, and Q. Lin, J. Cdl. Biochm., 17C, 301 (1993).