67 / 120
Liposomes, Exosomes, and Virosomes: From Modeling Complex
Membrane Processes to Medical Diagnostics and Drug Delivery
Poster Abstracts
62
31-POS
Board 16
PTEN and VSPs: On the Way to Identify the Structural Origin for Their Substrate
Specificity
Kirstin Hobiger
1
, Michael G. Leitner
1
, Tillmann Utesch
2
, Anja Feuer
1
, Maria Andrea
Mroginski
2
, Dominik Oliver
1
, Christian R. Halaszovich
1
.
2
Technische Universität Berlin, Berlin, Germany.
1
Philipps-Universität Marburg, Marburg,
Germany,
The phosphatase and tensin homolog deleted on chromosome ten (PTEN) is one of the most
crucial tumor suppressor proteins in mammals. It counteracts the PI3-kinase signaling cascade by
cleaving 3’-site phosphate groups from membranous phosphoinositides (PIs). In doing so, PTEN
prevents unlimited cell proliferation and tumor genesis. The phosphatase is a soluble protein that
transiently binds to the membrane surface for catalysis. This property hardens the experimental
access to PTEN's activity for direct interventions. Voltage-sensitive phosphatases (VSPs)
overcome this limitation. The enzymes share high sequential and structural homology to PTEN.
However, their enzymatic activity is coupled to the transmembrane potential, which allows a
reversible depletion of PIs under experimental control. By substituting the catalytic domain of
the VSP from
Ciona intestinalis
(Ci-VSP) against that one of PTEN, we created a voltage-
switchable VSP-chimera with the 3’-site PI-specificity of PTEN. Because VSPs prefer to
dephosphorylate the 5’-site position of PI-substrates, we are currently studying native VSPs in
comparison to the VSP-chimera to identify the structural origin for the substrate specificity of the
phosphatases. For this purpose, we use an approach that combines molecular dynamics
simulations and phosphatase activity assays in mammalian cells and in vitro. First experiments
revealed the importance of the membrane environment for the regulation of the substrate
specificity. In particular, the reaction toward one PI-substrate seems to be controlled by an
allosteric mechanism that is mediated by at least one of the flexible loops that surround the active
site. Since this regulation process depends on the properties of the membrane used for the
phosphatase assay, we are currently searching for the optimal membrane system that will help us
to identify the structural determinants for the substrate specificity of PTEN and its homologs.