A novel partial agonist of the exchange protein activated by cAMP isoform 1
(EPAC1), I942, was recently discovered and shown to reduce the guanine exchange
factor activity of cAMP-bound EPAC1 to approximately 10% relative to cAMP
activation. However, the inhibition mechanism of I942 remains unknown. Here, we
utilize NMR spectroscopy to probe the inhibitory I942 - EPAC1 interactions at atomic
resolution. The EPAC1 - I942 interface was mapped through intermolecular NOEs
measured by 15N and 13C filtered NOESY-HSQC experiment. Intermolecular NOE
mapping combined with other protein NMR methods, such as saturation transfer
difference, transfer Nuclear Overhauser Effect spectroscopy and chemical shift mapping,
we revealed that I942 interacts with the phosphate binding cassette (PBC) and base
binding region (BBR) of the EPAC1 cyclic nucleotide binding (CNB) domain, similar to
cAMP. The PBC controls the conformation of the hinge region, and subsequently,
allosterically shifts the hinge region between its active/inactive states. Molecular
dynamics simulation based on the NMR spectroscopy data revealed that EPAC1-CNB
adopts an intermediate conformation between its inactive and active states, which
explains the partial agonist nature of I942. / Thesis / Master of Science (MSc) / The exchange protein activated by cAMP (EPAC) is a receptor for the classical
secondary messenger cAMP. EPAC is present in multiple human systems and plays a
pivotal role in the development of a wide range of diseases. In this study, we aim to
establish the inhibition mechanism of a novel small molecule EPAC inhibitor/partial
agonist I942 using NMR spectroscopy with the goal of achieving a better understanding
of EPAC inhibition and paving the way for new small molecule EPAC inhibitors that can
potentially treat EPAC-related diseases such as heart failure and diabetes.
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/24806 |
| Date | January 2019 |
| Creators | Shao, Hongzhao |
| Contributors | Melacini, Giuseppe, Chemistry and Chemical Biology |
| Source Sets | McMaster University |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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