Out-licensing

Expression of Human Glutaminyl Cyclase in an Escherichia coli System: Structural Analysis and Inhibitor Designs

Introduction/Background

Glutaminyl cyclase is a catalyst responsible for protein N-terminal pyroglutamate formation. This enzyme is very useful in the commercially large-scale production of bioactive hormones and anticancer drugs which need the N-terminal pyroglutamate residue for their biological activities. In addition, human glutaminyl cyclase is a possible drug target for the treatment of several human disorders, such as Alzheimer's disease.

Aims/Hypothesis

We aim to express human glutaminyl cyclase in an Escherichia coli system.

Research

We have cloned glutaminyl cyclase cDNA from human bone marrow cDNA library. The protein was expressed in Escherichia coli system with the yields higher than approximately 10 mg/L bacterial culture, using a thioredoxin-tagged expression vector with several modifications. Based on high histidine content ( approximately 5%) of the protein, a convenient purification step by Ni-affinity chromatography was designed, leading to near homogeneity of the purified human glutaminyl cyclase. The identity of the recombinant human glutaminyl cyclase was confirmed by mass spectrometry and circular dichroism spectroscopy. The enzyme was active on both synthetic and physiological substrates, and the activity could be inhibited by several imidazole, triazole, and tetrazole derivatives. An atomic absorption analysis demonstrated that human glutaminyl cyclase contains one zinc ion per protein molecule. We also obtained the human glutaminyl cyclase crystals, which belong to cubic, tetragonal, and rhombohedral forms. Our works are useful to acquire new insights into human and animal glutaminyl cyclase, particularly for future structural analysis and inhibitor designs.

Conclusion

Our expression system has a relatively higher level of the recombinant human glutaminyl cyclase. We provide a convenient method for separation and purification of human glutaminyl cyclase from Escherichia coli cell lysates.

Relevance/Opportunity

Commercially, glutaminyl cyclase may facilitate the large-scale production of clinically useful hormones and anticancer drugs in cases of that the N-terminal pyroglutamate is required in the biological functions of these proteins. Please enquire quoting reference no. 11A-940825 if you are interested in forming licensing or codevelopment partnerships.

Early Stage

US pending, TW pending