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Novel substrates of Mycobacterium tuberculosis PknH Ser/Thr kinase

Xingji Zheng1, K G Papavinasasundaram, Yossef Av-Gay

  • 1Department of Medicine, Division of Infectious Diseases, University of British Columbia, 2733 Heather St., Vancouver, BC, Canada V5Z 3J5.

Biochemical and Biophysical Research Communications
|February 9, 2007
PubMed

Insights

Mycobacterium tuberculosis PknH kinase phosphorylates new substrates, Rv0681 and DacB1, crucial for bacterial growth. These findings reveal novel signaling pathways involving PknH in tuberculosis.

Area of Science:

  • Microbiology
  • Biochemistry
  • Molecular Biology

Background:

  • PknH is a Ser/Thr protein kinase in Mycobacterium tuberculosis.
  • PknH regulates cell wall enzyme expression and in vivo growth.
  • Identifying PknH substrates is key to understanding its regulatory roles.

Purpose of the Study:

  • To identify and characterize novel substrates of PknH.
  • To investigate the role of PknH in new signaling pathways.
  • To understand the phosphorylation kinetics of PknH.

Main Methods:

  • Bioinformatic analysis to predict potential PknH substrates.
  • In vitro kinase assays using recombinant PknH.
  • Point mutation studies to confirm phosphorylation sites.
  • Kinetic studies (Vmax) to compare phosphorylation rates.

Main Results:

  • Identified 40 potential PknH substrates using bioinformatics.
  • Confirmed phosphorylation of two new substrates (Rv0681, DacB1) by PknH in vitro.
  • Verified phosphorylation at in silico-predicted sites via point mutation.
  • Demonstrated similar relative phosphorylation rates for Rv0681, DacB1, and EmbR by PknH.
  • Observed that Rv0681 and DacB1 lack an FHA domain, unlike EmbR.

Conclusions:

  • Rv0681 and DacB1 are novel substrates of PknH in Mycobacterium tuberculosis.
  • These substrates may participate in previously unknown PknH-mediated signaling pathways.
  • PknH plays a significant role in regulating diverse cellular processes in M. tuberculosis.

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