Dataset Information


Prokaryotic expression, purification and characterization of human cyclooxygenase-2.

ABSTRACT: Cyclooxygenase-2 (COX-2) is a key enzyme which catalyzes the conversion of arachidonic acid (AA) into prostaglandins (PGs). It plays an important role in pathophysiological processes, such as tumorigenesis, angiogenesis, inflammation and tumor cell drug resistance. Therefore, COX-2 has been viewed as an important target for cancer therapy. The preparation of COX-2 protein is an important initial step for the subsequent development of COX-2 inhibitors. In this study, we report a strategy to heterologously express truncated human COX-2 (trCOX-2) in Escherichia coli (E. coli) BL21(DE3) host cells. Following denaturation, purification and renaturation, we successfully obtained enzymatically active trCOX-2 containing 257 residues of the C-terminus. Homology modeling and molecular docking analyses revealed that trCOX-2 retained the predicted 3D catalytic domain structure and AA could still bind to its hydrophobic groove. Western blot analysis and ELISA indicated that the trCOX-2 still retained its characteristic antigenicity and binding activity, while COX assays revealed that trCOX-2 maintained its enzyme activity. On the whole, in this study, we provided a novel method to isolate trCOX-2 possessing AA binding and catalytic activities. This study thus lays a foundation to facilitate further investigations of COX-2 and offers a valuable method with which to achieve the prokaryotic expression of a eukaryotic membrane protein.


PROVIDER: S-EPMC5466394 | BioStudies | 2017-01-01

REPOSITORIES: biostudies

Similar Datasets

2012-01-01 | S-EPMC3297362 | BioStudies
1000-01-01 | S-EPMC4880957 | BioStudies
2019-01-01 | S-EPMC6692340 | BioStudies
2015-01-01 | S-EPMC4367288 | BioStudies
2011-01-01 | S-EPMC3172888 | BioStudies
2010-01-01 | S-EPMC2806742 | BioStudies
2014-01-01 | S-EPMC3967765 | BioStudies
2012-01-01 | S-EPMC3220805 | BioStudies
2016-01-01 | S-EPMC5024554 | BioStudies
2019-01-01 | S-EPMC6769623 | BioStudies