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. 1992 Feb;61(2):381–391. doi: 10.1016/S0006-3495(92)81844-1

More than two pyrrole tautomers of mesoporphyrin stabilized by a protein. High resolution optical spectroscopic study.

J Fidy 1, J M Vanderkooi 1, J Zollfrank 1, J Friedrich 1
PMCID: PMC1260254  PMID: 1547326

Abstract

Mesoporphyrin IX substituted horseradish peroxidase was studied by fluorescence line narrowing and hole burning techniques at cryogenic temperatures. The spectral data reveal that four pyrrole tautomeric configurations of the chromophore are populated within the protein under the influence of irradiation and/or thermal treatment, and the existence of a fifth and a sixth tautomeric configuration is also likely. The relative population of the tautomers changes upon deuterium substitution through modification of the phototransition rate, and also depends on pH, which changes the protonation of neighboring amino acids in the heme pocket. The energy separation of the origins of the tautomers is approximately 100 cm-1. The distribution of barrier heights separating the different tautomeric forms in the ground state and their distribution was determined by temperature cycling hole burning. The distributions can be approximated by Gaussians. The experiment directly yields the distributions on a relative temperature scale, and a model is presented to transform the barrier heights into energy values. It is suggested that the energies for the tautomers are split partially due to the protein crystal field and that the trapping of the tautomeric forms is the consequence of interactions with neighboring amino acids within the heme pocket.

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Selected References

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