β-Amino acid substitution to investigate the recognition of angiotensin II (AngII) by angiotensin converting enzyme 2 (ACE2).
In spite of the important role of angiotensin converting enzyme 2 (ACE2) in the cardiovascular system, little is known about the substrate structural requirements of the AngII-ACE2 interaction. Here we investigate how changes in angiotensin II (AngII) structure affect binding and cleavage by ACE2. A series of C3 β-amino acid AngII analogs were generated and their secondary structure, ACE2 inhibition, and proteolytic stability assessed by circular dichroism (CD), quenched fluorescence substrate (QFS) assay, and LC-MS analysis, respectively. The β-amino acid-substituted AngII analogs showed differences in secondary structure, ACE2 binding and proteolytic stability. In particular, three different subsets of structure-activity profiles were observed corresponding to substitutions in the N-terminus, the central region and the C-terminal region of AngII. The results show that β-substitution can dramatically alter the structure of AngII and changes in structure correlated with ACE2 inhibition and/or substrate cleavage. β-amino acid substitution in the N-terminal region of AngII caused little change in structure or substrate cleavage, while substitution in the central region of AngII lead to increased β-turn structure and enhanced substrate cleavage. β-amino acid substitution in the C-terminal region significantly diminished both secondary structure and proteolytic processing by ACE2. The β-AngII analogs with enhanced or decreased proteolytic stability have potential application for therapeutic intervention in cardiovascular disease.
Clayton
Daniel
D
Biochemistry & Molecular Biology, Monash University, Clayton, Victoria 3800, Australia.
Hanchapola
Iresha
I
Hausler
Nicholas
N
Unabia
Sharon
S
Lew
Rebecca A
RA
Widdop
Robert E
RE
Smith
Alexander I
AI
Perlmutter
Patrick
P
Aguilar
Marie-Isabel
MI
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000804:Angiotensin II / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D000085962:Angiotensin-Converting Enzyme 2; D002942:Circular Dichroism; D008969:Molecular Sequence Data; D007703:Peptidyl-Dipeptidase A / Q000378:metabolism*
D000804:Angiotensin II; D007703:Peptidyl-Dipeptidase A; D000085962:Angiotensin-Converting Enzyme 2