Functionally important structural elements of the cyanobacterial clock-related protein Pex

Shunsuke Kurosawa, Reiko Murakami, Kiyoshi Onai, Megumi Morishita, Daisuke Hasegawa, Ryo Iwase, Tatsuya Uzumaki, Fumio Hayashi, Tomomi Kitajima-ihara, Shuhei Sakata, Midori Murakami, Tsutomu Kouyama, Masahiro Ishiura

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4 Citations (Scopus)

Abstract

Pex, a clock-related protein involved in the input pathway of the cyanobacterial circadian clock system, suppresses the expression of clock gene kaiA and lengthens the circadian period. Here, we determined the crystal structure of Anabaena Pex (Ana Pex; Anabaena sp. strain PCC 7120) and Synechococcus Pex (Syn Pex; S ynechococcus sp. strain PCC 7942). Pex is a homodimer that forms a winged-helix structure. Using the DNase I protection and electrophoresis mobility shift assays on a Synechococcus kaiA upstream region, we identified a minimal 25-bp sequence that contained an imperfectly inverted repeat sequence as the Pex-binding sequence. Based on crystal structure, we predicted the amino acid residues essential for Pex's DNA-binding activity and examined the effects of various Ala-substitutions in the α3 helix and wing region of Pex on in vitro DNA-binding activity and in vivo rhythm functions. Mutant Ana Pex proteins carrying a substitution in the wing region displayed no specific DNA-binding activity, whereas those carrying a substitution in the α3 helix did display specific binding activity. But the latter were less thermostable than wild-type Ana Pex and their in vitro functions were defective. We concluded that Pex binds a kaiA upstream DNA sequence via its wing region and that its α3 helix is probably important to its stability.

Original languageEnglish
Pages (from-to)1-16
Number of pages16
JournalGenes to Cells
Volume14
Issue number1
DOIs
Publication statusPublished - 2009 Jan 2
Externally publishedYes

ASJC Scopus subject areas

  • Genetics
  • Cell Biology

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    Kurosawa, S., Murakami, R., Onai, K., Morishita, M., Hasegawa, D., Iwase, R., Uzumaki, T., Hayashi, F., Kitajima-ihara, T., Sakata, S., Murakami, M., Kouyama, T., & Ishiura, M. (2009). Functionally important structural elements of the cyanobacterial clock-related protein Pex. Genes to Cells, 14(1), 1-16. https://doi.org/10.1111/j.1365-2443.2008.01245.x