Recruitment of governing elements for eletron transfer in the nitric oxide synthase family

M. Jáchymová, P. Martásek, S. Panda, L. J. Roman, M. Panda, T. M. Shea, Y. Ishimura, J. J.P. Kim, B. S.S. Masters

Research output: Contribution to journalArticle

31 Scopus citations

Abstract

At least three building blocks are responsible for the molecular basis of the modulation of electron transfer in nitric oxide synthase (NOS) isoforms: the calmodulin-binding sequence, the C-terminal extension, and the autoregulatory loop in the reductase domain. We have attempted to impart the control conferred by the C termini of NOS to cytochrome P450 oxidoreductase (CYPOR), which contains none of these regulatory elements. The effect of these C termini on the properties of CYPOR sheds light on the possible evolutionary origin of NOS and addresses the recruitment of new peptides on the development of new functions for CYPOR. The C termini of NOSs modulate flavoprotein-mediated electron transfer to various electron acceptors. The reduction of the artificial electron acceptors cytochrome c, 2,6-dichlorophenolindophenol, and ferricyanide was inhibited by the addition of any of these C termini to CYPOR, whereas the reduction of molecular O2 was increased. This suggests a shift in the rate-limiting step, indicating that the NOS C termini interrupt electron flux between flavin mononucleotide (FMM) and flavin adenine dinucleotide (FAD) and/or the electron acceptors. The modulation of CYPOR by the addition of the NOS C termini is also supported by flavin reoxidation and fluorescence-quenching studies and antibody recognition of the C-terminal extension. These experiments support the origin of the NOS enzymes from modules consisting of a heme domain and CYPOR or ferredoxin-NADP+ reductase- and flavodoxin-like subdomains that constitute CYPOR, followed by further recruitment of smaller modulating elements into the flavin-binding domains.

Original languageEnglish (US)
Pages (from-to)15833-15838
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume102
Issue number44
DOIs
StatePublished - Nov 1 2005

Keywords

  • Flavoprotein
  • NADPH-cytochrome P450 reductase
  • Protein evolution

ASJC Scopus subject areas

  • General

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    Jáchymová, M., Martásek, P., Panda, S., Roman, L. J., Panda, M., Shea, T. M., Ishimura, Y., Kim, J. J. P., & Masters, B. S. S. (2005). Recruitment of governing elements for eletron transfer in the nitric oxide synthase family. Proceedings of the National Academy of Sciences of the United States of America, 102(44), 15833-15838. https://doi.org/10.1073/pnas.0506522102