Novel Variants of Angiotensin Converting Enzyme-2 of Shorter Molecular Size to Target the Kidney Renin Angiotensin System

Wysocki, Jan; Schulze, Arndt; Batlle, Daniel

Enzyme activity in the conditioned culture media, from HEK cells transiently transfected with the ACE2 constructs generated through truncation of the C terminus (1-619 AA and 1-605 AA), was at least as high as in the media collected from cells overexpressing the native rACE2 1-740, as judged by the kinetic slope ( Figure 4 ). In contrast, transfection with a shorter ACE2 fragment (1-522 AA) did not show any enzyme activity ( Figure 4A ). The two enzymatically active short ACE2 variants (619 and 605) appeared as a single band at the apparent molecular weight of~70 kDa, as expected from the amino acid sequence deducted using the Expasy Informatics tool ( Figure 4B and Figure S3 ). The enzymatically inactive variant 1-522 appeared, in addition to the expected size of~50-60 kD, in oligomerized forms of 2-3 bands of higher molecular weight ( Figure 4B ). This suggests that the minimal length requirement on the C-terminal end for murine ACE2 to be enzymatically active is contained between AA 522 and 605.