Structural and functional significance of arginine at position 2 of angiotensin II: Implications for oxidant-mediated amino acid residue conversions.

Angiotensin II (Ang II) is a peptide hormone that regulates vascular tone and blood pressure. Angiotensin-converting enzyme generates Ang II from angiotensin I, whereas angiotensin-converting enzyme 2 (ACE2) cleaves Ang II to angiotensin 1-7. We previously showed that the arginine residue at the second position (Arg2) of Ang II is critical for its interaction with ACE2 as a substrate. The present study examined the mechanistic role of Arg2 in Ang II recognition by ACE2 and the functional consequences of its modification. Cleavage of Arg2 from Ang II, or replacement of Arg2 with other amino acids, eliminated Ang II's ability to compete with the fluorogenic substrate in the ACE2 activity assay. The Arg2-to-Glu substitution also affected the interactions of angiotensin I and angiotensin III with ACE2, confirming Arg2 as a critical determinant of substrate recognition by the enzyme. Since oxidation of Arg and Pro residues generates glutamic semialdehyde, which can be further oxidized to Glu, we previously proposed that reactive oxygen species promote oxidant-mediated amino acid conversion among Arg, Pro, and Glu residues. Thus, Arg2 of Ang II could undergo oxidant-mediated amino acid residue conversion to either Glu or Pro. Mass spectrometry confirmed the presence of both circulating Ang II (Arg2Glu) and Ang II (Arg2Pro) in rats. Global gene expression analyses further revealed that Arg2Glu- and Arg2Pro-converted Ang II peptides elicit cellular signaling responses distinct from those induced by wild-type Ang II, suggesting that oxidant-mediated modifications of Arg2 generate naturally occurring angiotensin peptides with unique biological functions.