FMP Publications

Our publications are recorded in a searchable database since 2010, updates will be added regularly.

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Different inhibition of Gbetagamma-stimulated class IB phosphoinositide 3-kinase (PI3K) variants by a monoclonal antibody. Specific function of p101 as a Gbetagamma-dependent regulator of PI3Kgamma enzymatic activity
Shymanets(*), A., Vadas(*), O., Czupalla(*), C., LoPiccolo(*), J., Brenowitz(*), M., Ghigo(*), A., Hirsch(*), E., Krause, E., Wetzker(*), R., Williams(*), R. L., Harteneck(*), C.; Nürnberg(*), B.
Biochem J, 469:59-69

Tags: Mass Spectrometry (Krause, E.)

Abstract: Class IB phosphoinositide 3-kinases gamma (PI3Kgamma) are second-messenger-generating enzymes downstream of signalling cascades triggered by G-protein-coupled receptors (GPCRs). PI3Kgamma variants have one catalytic p110gamma subunit that can form two different heterodimers by binding to one of a pair of non-catalytic subunits, p87 or p101. Growing experimental data argue for a different regulation of p87-p110gamma and p101-p110gamma allowing integration into distinct signalling pathways. Pharmacological tools enabling distinct modulation of the two variants are missing. The ability of an anti-p110gamma monoclonal antibody [mAb(A)p110gamma] to block PI3Kgamma enzymatic activity attracted us to characterize this tool in detail using purified proteins. In order to get insight into the antibody-p110gamma interface, hydrogen-deuterium exchange coupled to MS (HDX-MS) measurements were performed demonstrating binding of the monoclonal antibody to the C2 domain in p110gamma, which was accompanied by conformational changes in the helical domain harbouring the Gbetagamma-binding site. We then studied the modulation of phospholipid vesicles association of PI3Kgamma by the antibody. p87-p110gamma showed a significantly reduced Gbetagamma-mediated phospholipid recruitment as compared with p101-p110gamma. Concomitantly, in the presence of mAb(A)p110gamma, Gbetagamma did not bind to p87-p110gamma. These data correlated with the ability of the antibody to block Gbetagamma-stimulated lipid kinase activity of p87-p110gamma 30-fold more potently than p101-p110gamma. Our data argue for differential regulatory functions of the non-catalytic subunits and a specific Gbetagamma-dependent regulation of p101 in PI3Kgamma activation. In this scenario, we consider the antibody as a valuable tool to dissect the distinct roles of the two PI3Kgamma variants downstream of GPCRs.

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Leibniz-Forschungsinstitut für Molekulare Pharmakologie im Forschungsverbund Berlin e.V. (FMP)
Campus Berlin-Buch
Robert-Roessle-Str. 10
13125 Berlin, Germany
+4930 94793 - 100 
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