The conserved active site histidine-glutamate pair of ferrochelatase coordinately catalyzes porphyrin metalation

Research output: Contribution to journalArticle

Authors

  • Gregory A. Hunter
  • Sai Lakshmana Vankayala
  • Mallory E. Gillam
  • Fiona L. Kearns
  • H. Lee Woodcock
  • Gloria C. Ferreira

External Institution(s)

  • University of South Florida

Details

Original languageEnglish (US)
Pages (from-to)556-569
Number of pages14
JournalJournal of Porphyrins and Phthalocyanines
Volume20
Issue number1-4
StatusPublished - Apr 1 2016
Peer-reviewedYes

Abstract

Ferrochelatase catalyzes the insertion of ferrous iron into protoporphyrin IX to generate heme. Despite recent research on the reaction mechanism of ferrochelatase, the precise roles and localization of individual active site residues in catalysis, particularly those involved in the insertion of the ferrous iron into the protoporphyrin IX substrate, remain controversial. One outstanding question is from which side of the macrocycle of the bound porphyin substrate is the ferrous iron substrate inserted. Pre-steady state kinetic experiments done under single-turnover conditions conclusively demonstrate that metal ion insertion is pH-dependent, and that the conserved active site His-Glu pair coordinately catalyzes the metal ion insertion reaction. Further, pKa calculations and molecular dynamic simulations indicate that the active site His is deprotonated and the protonation state of the Glu relates to the conformational state of ferrochelatase. Specifically, the conserved Glu in the open conformation of ferrochelatase is deprotonated, while it remains protonated in the closed conformation. These findings support not only the role of the His-Glu pair in catalyzing metal ion insertion, as these residues need to be deprotonated to bind the incoming metal ion, but also the importance of the relationship between the protonation state of the Glu residue and the conformation of ferrochelatase. Finally, the results of this study are consistent with our previous proposal that the unwinding of the π-helix, the major structural determinant of the closed to open conformational transition in ferrochelatase, is associated with the Glu residue binding the Fe2+ substrate from a mitochondrial Fe2+ donor.

    Research areas

  • CHARMM, PropKa, chelatase, enzyme, ferrochelatase, iron, molecular dynamic simulations, p K a, porphyrin, porphyrin metalation, protoporphyrin IX, tetrapyrrole

Citation formats

APA

Hunter, G. A., Vankayala, S. L., Gillam, M. E., Kearns, F. L., Lee Woodcock, H., & Ferreira, G. C. (2016). The conserved active site histidine-glutamate pair of ferrochelatase coordinately catalyzes porphyrin metalation. Journal of Porphyrins and Phthalocyanines, 20(1-4), 556-569. https://doi.org/10.1142/S1088424616500395

Harvard

Hunter, GA, Vankayala, SL, Gillam, ME, Kearns, FL, Lee Woodcock, H & Ferreira, GC 2016, 'The conserved active site histidine-glutamate pair of ferrochelatase coordinately catalyzes porphyrin metalation', Journal of Porphyrins and Phthalocyanines, vol. 20, no. 1-4, pp. 556-569. https://doi.org/10.1142/S1088424616500395