Project description:We use HDX-MS to interrogate the AKT1 DrLink conformational changes upon binding AKT1 active site inhibitors A-443654, Capivasertib, and Uprosertib, Akt1 allosteric inhibitor MK-2206, and ADP.
Project description:The class IB phosphoinositide 3-kinase (PI3K), PI3K, is a master regulator of immune cell function, and is a promising drug target for both inflammatory diseases and cancer. Critical to PI3K function is the association of the p110 catalytic subunit to either a p101 or p84 regulatory subunit, which mediates regulation by G-protein coupled receptors (GPCRs). Here, we report the first structure of a heterodimeric PI3K complex, p110-p101. This structure revealed a unique mode of assembly of catalytic and regulatory subunits distinct from that of other class I PI3K complexes. Multiple oncogenic mutations mapped to these novel interfaces led to increased activation by G. p101 mediates activation through its G binding domain, recruiting the complex to the membrane and allowing for engagement of a secondary site in p110. A nanobody that specifically binds to this p101-G interface blocks activation providing a novel tool to study p101-specific signaling events in vivo.
Project description:We used HDXMS to study the confomational changes induced in a PIKfyve construct upon binding PI3P membrane and a variety of inhibitors. Here we showed a significant protection in deuterium exchange upon binding all inhibitors, and regions of protection and exposure upon binding PI3P membrane.
Project description:To examine the interaction between PI4KIIIa and Calcineurin, HDX-MS experiments comparing calcineurin or PI4KA FAM delta C alone to the calcineurin and PI4KA FAM delta C complex were carried out.
Project description:Hydrogen deuterium exchange mass spectrometry (HDX-MS) was used to analyze how the mng-SHIP1-dCT construct interacts receptor-derived phosphotryrosine peptides (pY) to probe how these peptides regulate SHIP1 autoinhibition. This HDX-MS helped to identify intramolecular contacts involved in the regulation of SHIP1 autoinibition. Results from this HDX analyzing the dimeric mng-SHIP1 construct are used in tandem with previous results analyzing the monomeric mini-SHIP construct to confirm the same mechanism of autoinhibtion exists. These results help confirm autoinhibition of monomeric SHIP1 is indeed likely regulated by the same mechanism to that of dimeric SHIP1.
Project description:Hydrogen deuterium exchange mass spectrometry of PLIN3 in the presence of three different membrane vesicles to analyze structural changes induced by membrane binding.