New fluorescent method for the histochemical detection of tripeptidyl peptidase I using glycyl-l-prolyl-l-met-2-anthraquinonyl hydrazide as substrate.
A new substrate for tripeptidyl peptidase I (TPP I; E.C.3.4.14.9)-Gly-L-Pro-L-Met-2-anthraquinonyl hydrazide (Gly-Pro-Met-2-AH) is synthesized and used for the fluorescent histochemical detection of the enzyme. The enzyme liberates low soluble 2-anthraquinonylhydrazine, which-couples quickly with 3-nitrobenzaldehyde (3-NBA) yielding a highly fluorescent water-insoluble hydrazone--3-nitrobenzylidene-2-anthraquinonylhydrazone. The latter compound is localized precisely at sites of enzymatic activity and marks them with a very bright and stable orange-red fluorescence after excitation with conventional monochromatic andlaser green light (lambda(exc)=520-580 nm). The new technique is used successfully for the visualization of the enzyme in tissue sections of different rat organs - and represent the first fluorescent histochemical method for that peptidase.
Dikov
A
A
Institute of Anatomy/Anatomy II, Friedrich Schiller University Jena, 07743 Jena, Germany.
Dimitrova
M
M
Krieg
R
R
Halbhuber
K J
KJ
D000626:Aminopeptidases; D000818:Animals; D000880:Anthraquinones / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D004151:Dipeptides / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism*; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D010450:Endopeptidases / Q000032:analysis*; D005260:Female; D005456:Fluorescent Dyes / Q000737:chemistry; D006651:Histocytochemistry / Q000379:methods*; D008297:Male; D008856:Microscopy, Fluorescence / Q000379:methods*; D051381:Rats; D017208:Rats, Wistar; D057057:Serine Proteases; D013379:Substrate Specificity; D000091346:Tripeptidyl-Peptidase 1
D000880:Anthraquinones; D004151:Dipeptides; D005456:Fluorescent Dyes; C000717269:Tpp1 protein, rat; D000091346:Tripeptidyl-Peptidase 1; C502018:glycyl-l-prolyl-l-met-2-anthraquinonylhydrazide; D010450:Endopeptidases; D057057:Serine Proteases; D000626:Aminopeptidases; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases