Abstract
Depression is associated with changes in cyclic guanosine monophosphate (cGMP) levels. Depression can be improved by increasing the cGMP concentration through the cGMP/PKG pathway with PDE2A inhibitors. This study is aimed to improve the expression of a highly active PDE2A protein with an Escherichia coli vector ST6 for the screening of PDE2A inhibitors. PDE2A gene was obtained through polymerase chain reaction. A recombinant plasmid of ST6-PDE2A was built by seamless cloning and then introduced into E. coli BL21 (DE3). The cultivation conditions were optimized to increase target protein expression. The expressed protein was purified with Ni-NTA affinity chromatography. Its purified activity was measured by a PDE-GloTM phosphodiesterase kit. An maximized protein expression was obtained by cultivating E. coli BL21 with ST6-PDE2A in the YT medium at 37 °C till OD600 reached to 0.6-0.8 and then by inducible expressing with 1 mM IPTG at 16 °C for 40 hours. The resultant active protein has an EC50 of 0.1196 mg/ml.
Highlights
Depression is a common mood and emotional disorder
Depression can be improved by increasing the cyclic guanosine monophosphate (cGMP) concentration through the cGMP/PKG pathway with PDE2A inhibitors
An maximized protein expression was obtained by cultivating E. coli BL21 with ST6-PDE2A in the YT medium at 37 °C till OD600 reached to 0.6-0.8 and by inducible expressing with 1 mM IPTG at 16 °C for 40 hours
Summary
Statistics from Worth Health Organization suggest that globally, the rate of depression is 11% and there are up to 1 million suicides associated with depression every year (Mann JJ, 2003; Song Y et a1., 2008) This disease is manifested as persistent and significant low mood, loss of interest, and low self-esteem. Phosphodiesterases (PDEs), which mainly act on cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), have recently attracted attention from the field of depression therapy. Both cAMP and cGMP can regulate the corresponding downstream protein kinases.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
