Anastasiia Bovdilova - AG Maurino

 

Post-translational regulation and structural characterization of major C4 proteins    

The photosynthetic isoform of NADP-malic enzyme (NADP-ME) in maize belongs to the most important and well-studied C4 enzymes. This C4 enzyme is thought to have evolved from a non-C4 chloroplastic ancestor. As known for some other well-studied C4 proteins (e.g. pyruvate, orthophosphate dikinase), one important molecular mechanism, which may be involved in protein regulation, is post-translational modifications (PTM). Until now, no information is available concerning the participation of PTMs in the regulation of the C4 NADP-ME.

Recently, one putative phosphorylation site in the maize photosynthetic NADP-ME has been found. We first have assessed the significance of this site by performing site-directed mutagenesis of the wild-type NADP-ME to mimic the phosphorylated state. The mutated protein was then heterologously produced in E. coli cells and purified by affinity chromatography.

The next step is determination of the enzymatic properties of mutated protein and their comparison to the corresponding features of the wild-type enzyme. The most important analysed properties are: pH optimum, kinetic parameters (Km and kcat values for L-malic acid and NADP), regulatory properties such as inhibition by malate and analysis of the enzyme’s structural integrity (verification of the protein conformation by circular dichroism, analytical ultracentrifugation and native PAGE).

We also investigate the functional importance of the PTMs in vivo by using antibodies specific for the phosphorylated and non-phosphorylated regions of the enzyme. This analysis allows to determine the time point(s) of the phosphorylation event(s) and its strength. This provides more deep insights into the role of the phosphorylation of the C4 NADP-ME in the physiology of maize.

 

Starting date: 01.03.2014

Thesis committee members: Dr. Veronica Maurino, Prof. Peter Westhoff

Mini Academic CV:

  • 2013: Master of Science at the Institute for Plant Biology and Biotechnology, Münster/Germany, group of Prof. Dr. B. Moerschbacher
    • Thesis: "Insights into priming activities of chitosan oligosaccharides produced using bacterial GH families 18 and 19 chitinases towards plant cells"
  • 2010: Bachelor of science in industrial biotechnology (at the National Technical University of Ukraine in Kiev, Kyiv Polytechnic Institute)

 

 

Photo of Anastasiia  Bovdilova

Anastasiia Bovdilova M.Sc.

Building: 26.03
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