Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.23.5 (cathepsin D)
 4,130 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The suitability of Z-Arg-Gly-Phe-Phe-Leu-MNA and Z-Arg-Gly-Phe-Phe-Pro-MNA for the assessment of cathepsin D activity was tested in biochemical and histochemical experiments. Substrates were dissolved in dimethylformamide and used at 0.1-0.5 mM in various buffers over a pH range of 3.5-7.4. Homogenates of various rat organs and isolated purified enzymes [cathepsin D from bovine spleen, dipeptidyl peptidase (DPP) IV from porcine kidney and rat lung] were used as enzyme sources. Pepstatin, di-isopropylfluorophosphate (DFP), p-chloromercuribenzoate, o-phenanthroline and a series of DPP IV inhibitors were used in inhibitor experiments. At pH 3.5 and 5.0, substrates were used in a two-step postcoupling procedure with aminopeptidase M and dipeptidyl peptidase IV as auxiliary enzymes and Fast Blue BB as coupling agent. Results were compared with those obtained with haemoglobin. Above pH 5.0 substrates were used in a one-step postcoupling procedure. Cryostat sections of snap-frozen or cold aldehyde-fixed tissue pieces of various rat organs and biopsies of human jejunal mucosa were used in histochemical experiments. As in biochemical tests a two-step procedure was used in the pH range 3.5-5.0, but Fast Blue B was used in the second step for the simultaneous coupling. Above pH 5.0 a one-step simultaneous azo coupling procedure was used with Fast Blue B as coupling agent. At pH 3.5 the hydrolysis rate of both synthetic substrates was about 100x lower than that of haemoglobin when cathepsin D from bovine spleen was used.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Are Z-Arg-Gly-Phe-Phe-Leu-MNA and Z-Arg-Gly-Phe-Phe-Pro-MNA suitable substrates for the demonstration of cathepsin D activity? 289 46

As a new approach, various synthetic fluorogenic substrates, the CellProbe reagents, were applied to examine the topography of their cleavage in vital human spermatozoa. These substrates are able to enter the cells without requiring previous cell permeabilization and can produce a fluorescent dye after cleavage, depending on enzyme activity. Vital spermatozoa from samples with normal spermiogram parameters showed fluorescence in different areas and intensity after incubation with a variety of substrates for aminopeptidase A, peroxides, subtilisin, dipeptidylpeptidase IV (DPP IV), cathepsin D, glucosidase and glucuronidase, but not with the substrate for galactosidase. Fluorescence was mainly located in the acrosomal cap (substrates for DPP IV, subtilisin, cathepsin D, glucosidase and glucuronidase) in the middle piece and head (substrates for peroxides, glucosidase), in the sperm head (substrates for aminopeptidase A) and occasionally in the tail (substrate for glucosidase). The substrate for subtilisin may play a role in andrology, because subtilisin is a serine protease like acrosin. This substrate may possibly be used to determine the acrosin activity in vital spermatozoa. The CellProbe reagents for fluorescence cytoenzymology may serve advanced methods in both clinical andrology and spermiological research, presuming that the characteristics and qualities of the synthetic substrates are correct. Therefore, more extended studies will be necessary to determine their clinical utility and significance under physiological and pathological conditions.
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PMID:Localisation of enzymes in live spermatozoa by CellProbe reagents (preliminary results). 994 87