Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P15088 (mast cell)
 14,925 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Various N-alpha-hydroxyalkyl derivatives of N-acyl amino acids and di- and tripeptides were prepared by hydrolysis or aminolysis of N-acyl 5-oxazolidinones. The stability of these derivatives was studied in aqueous solution as a function of pH. The compounds were all degraded quantitatively to their parent N-acylated amino acid or peptide and aldehyde but with vastly different rates. At pH 7.4 and 37 degrees C the half-lives of decomposition ranged from 4 min to 1500 hr. The structural factors influencing the stability included both steric and polar effects within the acyl and N-alpha-hydroxyalkyl moieties as well as within the amino acid attached to the N-alpha-hydroxyalkylated N-acyl amino acid. Whereas the N-benzyloxycarbonyl (Z) derivatives of the dipeptides Gly-L-Leu and Gly-L-Ala were readily hydrolyzed by carboxypeptidase A, the N-hydroxymethylated compounds, i.e., Z-Gly(CH2OH)-Leu and Z-Gly(CH2OH)-Ala, were resistant to cleavage by the enzyme as revealed by their similar rates of decomposition in the presence or absence of the enzyme at pH 7.4 and 37 degrees C. The results suggest that N-alpha-hydroxyalkylation of a peptide bond protects not only this bond but also an adjacent peptide bond against proteolytic cleavage. Since the N-alpha-hydroxyalkyl derivatives are readily bioreversible, undergoing spontaneous hydrolysis at physiological pH, this prodrug approach promises to overcome the enzymatic barrier to absorption of various peptides.
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PMID:Prodrugs of peptides. 9. Bioreversible N-alpha-hydroxyalkylation of the peptide bond to effect protection against carboxypeptidase or other proteolytic enzymes. 205 17

We tested four synthetic substances for their histochemical value to demonstrate the catalytic activities of chymase or tryptase in mast cells in sections of human gut. Both Suc-Ala-Ala-Phe-4 methoxy-2-naphthylamide (MNA) and N-acetyl-L-methionine-alpha-naphthyl ester (alpha-N-O-Met) reacted with chymase but not tryptase in mast cells. Conversely, D-Val-Leu-Arg-MNA and Z-Ala-Ala-Lys-MNA were hydrolyzed by mast cell tryptase but not chymase. These results were confirmed by use of two inhibitors of chymotrypsin-like activity, chymostatin and Z-Gly-Leu-Phe-chloromethyl ketone (CK) and two inhibitors of trypsin-like activity, Tos-Lys-CK and D-Val-Leu-Arg-CK. Excellent staining reactions were obtained on cryostat sections of unfixed or aldehyde-fixed tissues and on paraffin sections of Carnoy-fixed tissues. For chymase, however, Suc-Ala-Ala-Phe-MNA is preferred on cryostat sections because it is more specific. On paraffin sections alpha-N-O-Met is preferred because other cells are not then stained. For tryptase, Z-Ala-Ala-Lys-MNA was more selective and more specific and is the preferred general purpose substrate on cryostat sections of aldehyde-fixed tissues and for paraffin sections. D-Val-Leu-Arg-MNA is the preferred substrate for cryostat sections of unfixed tissue. Only a limited number of mast cells showed a reaction for chymase, and these occurred mainly in the submucosa. All mast cells, however, gave a reaction for tryptase, and we recommend the use of either substrate for this enzyme for routine detection of mast cells in human tissues. Double staining for the two main mast cell proteases is most conveniently undertaken on paraffin sections of Carnoy-fixed tissues using MNA substrates for tryptase and alpha-N-O-Met for chymase.
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PMID:Enzyme histochemical discrimination between tryptase and chymase in mast cells of human gut. 264 38

Three thioamide peptides in which the oxygen atom of the scissile peptide bond is replaced by sulfur (denoted by (= S)) were synthesized and found to be good, convenient substrates for carboxypeptidase A. The thioamide bond absorbs strongly in the ultraviolet region, and enzymatic hydrolysis is monitored easily using a continuously recording spectrophotometric assay. The reaction follows Michaelis-Menten kinetics with kcat values of 68, 9.0, and 3.7 sec-1 and Km values of 0.83, 0.81, and 0.53 mM for Z-Glu-Phe(= S)-Phe, Z-Gly-Ala(= S)-Phe, and Z-Phe(= S)-Phe, respectively. Activities of the thioamides and their oxygen amide analogs were determined with a series of metal-substituted carboxypeptidases. The Cd(II), Mn(II), Co(II), and Ni(II) enzymes exhibit 30%-35%, 60%-85%, 150%-190%, and 40%-55% of the Zn(II) enzyme activity with the amide substrates; this compares with 240%-970%, 0%-15%, 340%-840%, and 30%-140% of the Zn(II) activity, respectively, with the thioamides. The activity of the Cu(II) and Hg(II) enzymes is less than 3% toward all substrates. Cadmium, a thiophilic metal, yields an enzyme which is exceedingly active with the thioamides; the kcat/Km values are 2.4-9.7-fold higher than with Zn(II) carboxypeptidase. In contrast, Mn(II), which has a relatively low affinity for sulfur, yields an enzyme with correspondingly low activity toward the thioamides. The results are consistent with a mechanism for peptide bond hydrolysis in which the metal atom interacts with the substrate carbonyl atom during catalysis.
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PMID:Thioamide substrate probes of metal-substrate interactions in carboxypeptidase A catalysis. 380 99

A method has been designed for the assay of pancreatic carboxypeptidase A in blood serum. It uses Z-Gly-Phe as the substrate and fluorimetric determination of the released phenylalanine in an amino acid analyser, which yields a measure of free carboxypeptidase A. In addition, the sum (free carboxypeptidase A + procarboxypeptidase A) can be determined on a second portion preincubated with trypsin, which converts the proenzyme to the active form. Determinations made in fifteen healthy individuals showed the presence of a measurable concentration of free carboxypeptidase A. In acute pancreatitis, total carboxypeptidase A is raised. An increase in circulating proenzyme is observed in some cases. Data from 46 patients show a good correlation between total carboxypeptidase A, lipase and immunoreactive trypsin. Differential determination of procarboxypeptidase A and free carboxypeptidase A provides an interesting new tool for the diagnosis of pancreatic disorders.
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PMID:Determination of pancreatic carboxypeptidase A in human blood serum. 619 12

Carbobenzoxythioglycyl-L-phenylalanine [CbzNHCH2C(==S)Phe, Z-Glys-Phe] was synthesized as thioamide analogue of Z-Gly-Phe, a known substrate of carboxypeptidase A (CPA). By use of a ninhydrin-based assay and Z-Gly-Gly-Phe as the substrate, Z-Glys-Phe was shown to be a weak competitive inhibitor of CPA (Ki = 1.4 mM). The L isomer (but not the D) of Z-Glys-Phe proved to be a substrate for CPA (Km = 1.1 mM and kcat = 5.3 s-1 at pH 7.5), binding with comparable affinity to, but hydrolyzing at 10% the rate of, the oxo analogue Z-Gly-Phe. The CPA-catalyzed hydrolysis of Z-Glys-Phe was shown to involve only C-N bond cleavage, to give carbobenzoxythioglycine and phenylalanine.
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PMID:A thioamide substrate of carboxypeptidase A. 708 37

We examined three tissue samples from each of four cows with non-lesional skin, tissue samples from a cow with multiple cutaneous mast cell tumors, and samples from another cow in which mast cells were infiltrating multiple lymphosarcomas of the skin, for the presence of tryptase and chymase by enzyme cytochemical and immunohistological methods. The enzyme activities of tryptase and chymase were tested using N-carbobenzoxy-glycilglycil-L-arginine-2-naphthylamide (Z-Gly-Gly-Arg-NA) and naphthol-AS-D-chloroacetate (N-AS-D-CA) as substrates, respectively. Tryptase reactivity could be demonstrated in frozen and Carnoy-fixed paraffin sections. Chymase reactivity was seen in neither frozen nor paraffin sections of formalin- or Carnoy-fixed skin tissues. Antibody linkage with a polyclonal rabbit anti-human skin tryptase antibody was highly specific in bovine normal cutaneous, infiltrating, and tumor mast cells. More than 90% of the tumor mast cells were distinctly tryptase-positive. With alcian blue, only slightly more than 10% of the mast cells stained clearly positive and with methylene blue hardly any staining of mast cell granules could be demonstrated. No antibody labeling of mast cell granules in any of the tissue sections was detected by the use of rabbit anti-dog chymase antiserum. These results indicate that there is a striking antigenic similarity of bovine tryptase to its canine and human equivalents. The demonstration of tryptase is an important tool in confirming the diagnosis of undifferentiated mast cell tumors. In contrast to other species, chymase appears to be completely absent in bovine skin mast cells.
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PMID:Demonstration of tryptase in bovine cutaneous and tumor mast cells. 756 Aug 96