Recombinant Expression and Characterization of Human and Murine ACE2: Species-Specific Activation of the Alternative Renin-Angiotensin-System

Figure 3

Interference of human and murine ACE2 with the endogenous RAS in human plasma. Anticoagulated plasma samples were incubated at 37°C in the presence of 100 pg/mL recombinant human renin (a, b). rhACE2 or rmACE2 were added as indicated at a final plasma concentration of 5 μg/mL. RAS-Fingerprints were measured by LC-MS/MS as indicated in Section 2. Samples were incubated in the absence (a) or presence (b) of 10 μM of the ACE inhibitor Lisinopril. In this figure, the diameter of the spheres reflects the concentration of the respective peptide metabolite, which is also given in pg/mL next to each individual sphere. 0 pg/mL indicates concentrations below quantification limits, which are defined by a signal-to-noise ratio below 10. Furthermore, the amino acid sequence of each angiotensin metabolite is schematically given in brackets beside the corresponding sphere. The sequence annotation is based on the decapeptide Angiotensin I (1–10) which is N- or C-terminally cleaved by the indicated proteases. Proteases are illustrated by arrows connecting their substrate and product. AP: aminopeptidases; NEP: neutral endopeptidase; DAP: di-aminopeptidase; ACE: angiotensin-converting enzyme. (c) Indicated concentrations of recombinant human renin were added to Lisinopril-treated plasma samples. Control samples (black bars), rmACE2- (grey bars), and rhACE2- (white bars) treated samples were subjected to RAS-Fingerprinting, and resulting concentrations for Ang 1–10 (left) and Ang 1–9 (right) are given in pg/mL.