New fluorogenic substrates for the study of secondary specificity of prolyl oligopeptidase.
The secondary specificity of prolyl oligopeptidase (POP) has been studied by using a series of fluorogenic substrates containing the highly fluorescent 7-amino-4-methyl-2-quinolinone (AMeq) marker. The substrates were dipeptides of the general formula Z-X-Pro-NH-Meq, bearing amino acid residues with variable functional groups [Met, Lys(Boc), Lys, His, Ser, Leu, Glu(OMe), Glu, Cys(Bzl)] at the P2 position, and the tripeptide Z-Asn-Cys(Bzl)-Pro-NH-Meq. The kinetic parameters for their hydrolysis by porcine kidney POP were determined at lambda ex = 360 nm and lambda em = 430 nm. All the dipeptide substrates showed a high affinity to the enzyme and could be used for its fluorometric determination. The S2 binding subsite of POP can accommodate amino acid residues with a bulky side group, while it prefers a positively charged group (free Lys) instead of a negatively charged one (free Glu).
Noula
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C
Department of Chemistry, University of Athens, Greece.
Kokotos
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G
Barth
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T
Tzougraki
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C
D005456:Fluorescent Dyes; D009682:Magnetic Resonance Spectroscopy; D000085822:Prolyl Oligopeptidases; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000378:metabolism; D016339:Spectrometry, Mass, Fast Atom Bombardment; D013379:Substrate Specificity
D005456:Fluorescent Dyes; D012697:Serine Endopeptidases; D000085822:Prolyl Oligopeptidases