Characterization of PgPepO, a bacterial homologue of endothelin-converting enzyme-1.
PgPepO is a homologue of endothelin-converting enzyme-1 (ECE-1), with which it shares 31% identity. PgPepO was isolated from the periodontal pathogen Porphyromonas gingivalis. Recent studies have suggested a link between periodontal and cardiovascular disease, and several groups have suggested that bacterial and viral infections may contribute to the latter. P. gingivalis possesses the ability to invade, and multiply within, aortic endothelial cells and has been localized to atherosclerotic plaques. PgPepO was expressed and purified to homogeneity and we have begun detailed functional analysis, in terms of substrate preference and inhibitor specificity, in order to provide active-site comparisons with other members of the neprilysin (NEP)/ECE family. PgPepO possesses similar substrate specificity to ECE-1 and has been shown to cleave big endothelin-1 (big ET-1), big ET-2 and big ET-3, converting the substrates into their respective mature endothelin peptides. Substance P, angiotensin I, angiotensin II and neurotensin are all cleaved at multiple sites by PgPepO and the kinetics of these reactions have been compared. The potent vasoconstrictor urotensin II is not hydrolysed by PgPepO. Cleavage of bradykinin by PgPepO occurs at the Pro(7)-Phe(8) bond and is inhibited by the NEP and ECE-1 inhibitor phosphoramidon in a pH-dependent fashion (IC(50) =10 microM at pH 7.0) but not by thiorphan, an NEP-specific inhibitor. PgPepO activity is completely inhibited by EDTA. Characterization of this enzyme is important in elucidating possible links between periodontal pathogens and cardiovascular disorders such as atherosclerosis, and provides an opportunity to gain structural information on a bacterial protein with striking similarity to human ECE-1.
School of Biochemistry and Molecular Biology, University of Leeds, Leeds LS2 9JT, U.K. bmbjac@bmb.leeds.ac.uk
Julie A
JA
Carson
Toshihiro
T
Ansai
Shuji
S
Awano
Weixian
W
Yu
Tadamichi
T
Takehara
Anthony J
AJ
Turner
D000803:Angiotensin I / Q000378:metabolism; D000804:Angiotensin II / Q000378:metabolism; D016282:Aspartic Acid Endopeptidases / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000494:pharmacology*; D001920:Bradykinin / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D000072396:Endothelin-Converting Enzymes; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000378:metabolism; D006020:Glycopeptides / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D020128:Inhibitory Concentration 50; D008666:Metalloendopeptidases; D009496:Neurotensin / Q000378:metabolism; D016966:Porphyromonas gingivalis / Q000201:enzymology*; D013373:Substance P / Q000378:metabolism; D015244:Thiorphan / Q000378:metabolism
D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D006020:Glycopeptides; D000804:Angiotensin II; D013373:Substance P; D009496:Neurotensin; D000803:Angiotensin I; D015244:Thiorphan; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; C000606923:ECE1 protein, human; D000072396:Endothelin-Converting Enzymes; D001920:Bradykinin; C008890:phosphoramidon