Analysis of the Glycoprotein Properties and Glycan Structures of Chicken Angiotensin Converting Enzyme 2

Glycosylation of angiotensin converting enzyme 2 (ACE2), the principal receptor protein of severe scute respiratory syndrome coronavirus 2 (SARS-CoV-2), plays a crucial role in coronavirus infection. However, the precise host range and intermediate host of SARS-CoV-2 remain uncertain, and our knowledge on the glycosylation mechanism of animal ACE2 receptor interaction remains limited. Therefore, it is necessary to search for new information about the glycan structures of ACE2 protein in animals. In this study, we used high performance liquid chromatography (HPLC) and matrix assisted laser ionization time of flight mass spectrometry (MALDI-TOF-MS) to characterize the glycoprotein properties and glycan structures of recombinant ACE2 protein, which is produced in CHO cells, in chickens. Our analyses revealed that O-glycans were structurally diverse, consisting of Core 1, 2 and 3 type glycans. Moreover, chicken ACE2 almost completely occupied the Core 2 glycan (68.89%). Concerning N-glycosylation, chicken ACE2 contained complex glycan (94.59%) and high-mannose glycan (5.41%). Furthermore, the N-glycans of chicken ACE2 protein were largely occupied by galactosylation and sulfation. Overall, this study provides additional structural details on glycans, which will help researchers study the mechanisms underlying animal host ACE2 and the immune response, as well as knowledge for developing urgently needed disease biomarkers and remedies.