Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Choline acetyltransferase (Acetyl-CoA:choline O-acetyltransferase, EC 2.3.1.6, abbreviated ChAT), the biosynthetic enzyme for acetylcholine and acetylcholinesterase (EC 3.1.1.7, abbreviated AChE) are expressed in a human cholinergic neuroblastoma cell line, MC-IXC. We have shown that ChAT activity can be regulated in culture by retinoic acid, an active metabolite of vitamin A, and by sodium butyrate, an organic fatty acid. Optimal concentrations of these agents produce 4.3-fold and 1.6-fold increases in ChAT activity, respectively. The effects of retinoic acid are statistically significant after 24 h, whereas for sodium butyrate significant differences are seen only after 48 h. Since retinoic acid stimulation of ChAT activity was reversed only by trypsin treatment and not by removal of retinoic acid from the medium, this suggests that this agent may be acting at the level of the cell surface. Other differentiating conditions, such as culture in serum-free medium or addition of 1-2% dimethylsulfoxide did not increase ChAT activity. Acetylcholinesterase activity was shown to increase only in the presence of sodium butyrate, suggesting that retinoic acid and sodium butyrate may be acting via different pathways. Retinoic acid and sodium butyrate both seem to be permissive rather than instructive in regulating ChAT activity in that they are unable to induce ChAT expression de novo in cell lines which do not already express ChAT activity.
...
PMID:Stimulation of choline acetyltransferase activity by retinoic acid and sodium butyrate in a cultured human neuroblastoma. 292 23

The effects of t-butylhydroperoxide (tBHP), its alkoxyl radical (tBuO.) and its peroxyl radical (tBuOO.) in model systems and on red blood cells were studied. Glyceraldehyde-3-phosphate dehydrogenase was strongly inhibited by tBHP via a direct reaction of the hydroperoxide with an essential sulfhydryl group in the enzyme molecule. Several other enzymes were unaffected by tBHP. Alcohol dehydrogenase was strongly inhibited by tBuO. but was much less sensitive to tBuOO.. Lysozyme, lactate dehydrogenase and trypsin, on the other hand, were very sensitive to the peroxyl and not, or much less, to the alkoxyl radical, whereas acetylcholinesterase was very sensitive to both radicals. tBuOO. caused covalent binding of tryptophan, tyrosine, histidine and methionine to serum albumin. The corresponding alkoxyl radical was ineffective in this respect. Conversely, tBuO. caused peroxidation of linolenic acid, whereas tBuOO. did not. Incubation of human erythrocytes with tBHP caused lipid peroxidation and K+ leakage. Both effects were caused by tBHP-derived radicals generated in a reaction of the hydroperoxide with hemoglobin. With radical scavengers it was possible to dissociate tBHP-induced lipid peroxidation and K+ leakage, demonstrating that these two processes are not causally related. Experimental results indicate that tBuO. causes lipid peroxidation, whereas tBuOO. is responsible for K+ leakage.
...
PMID:Inhibition of enzymes and oxidative damage of red blood cells induced by t-butylhydroperoxide-derived radicals. 293 Jul 85

Human serum cholinesterase was digested with pepsin under conditions which left disulfide bonds intact. Peptides were isolated by high pressure liquid chromatography, and those containing disulfide bonds were identified by a color assay. Peptides were characterized by amino acid sequencing and composition analysis. Human serum cholinesterase contains 8 half-cystines in each subunit of 574 amino acids. Six of these form three internal disulfide bridges: between Cys65-Cys92, Cys252-Cys263, and Cys400-Cys519. A disulfide bond with Cys65 rather than Cys66 was inferred by homology with Torpedo acetylcholinesterase. Cys571 forms a disulfide bridge with Cys571 of an identical subunit. This interchain disulfide bridge is four amino acids from the carboxyl terminus. A peptide containing the interchain disulfide is readily cleaved from cholinesterase by trypsin (Lockridge, O., and La Du, B. N. (1982) J. Biol. Chem. 257, 12012-12018), suggesting that the carboxyl terminus is near the surface of the globular tetrameric protein. The disulfide bridges in human cholinesterase have exactly the same location as in Torpedo californica acetylcholinesterase. There is one potential free sulfhydryl in human cholinesterase at Cys66, but this sulfhydryl could not be alkylated. Comparison of human cholinesterase, and Torpedo and Drosophila acetylcholinesterases to the serine proteases suggests that the cholinesterases constitute a separate family of serine esterases, distinct from the trypsin family and from subtilisin.
...
PMID:Location of disulfide bonds within the sequence of human serum cholinesterase. 311 73

Acetylcholinesterase (AChE, EC 3.1.1.7) purified from the electric organ of eel possesses a protease activity resembling that of a neuropeptide processing enzyme. To examine whether any mammalian AChEs possess a similar protease activity, the enzyme was purified, 110,000-fold from foetal bovine serum. Purified serum AChE cleaved 2 synthetic peptide substrates in a manner resembling the combined actions of trypsin-like and carboxypeptidase B-like enzymes. A synthetic fragment of preproenkephalin A (residues 97-107) containing a complete methionine-enkephalin sequence was cleaved by serum AChE to yield free methionine-enkephalin. The carboxypeptidase action of AChE was weakly stimulated by the presence of 100 microM CoCl2 suggesting the requirement of a metal ion for complete activity. The results support the hypothesis that in many tissues AChE may act as a neuropeptide processing enzyme.
...
PMID:Serum acetylcholinesterase possesses trypsin-like and carboxypeptidase B-like activity. 322 17

Acetylcholinesterase (AChE) is one of the most highly studied enzymes, although its function in many tissues has remained obscure. AChE purified from eel or foetal bovine serum possesses proteolytic activity in addition to esterase activity. The presence of trypsin-like and metallocarboxypeptidase-like activities associated with AChE accounts for its ability to convert enkephalin peptide precursors into enkephalins. Several lines of evidence indicate that AChE's trypsin-like activity is an integral component of the molecule and that it is activated by autolysis. Incubation of affinity-purified eel AChE generated several fragments of low relative molecular mass (Mr). One of these low Mr fragments (Mr = 25,000 Da, 25K) cleaved from the 70K form of AChE, possessed considerable sequence similarity to the N-terminal sequence of pancreatic trypsin. Autolysis of eel AChE may give rise to a neuropeptide processing enzyme.
...
PMID:Acetylcholinesterase undergoes autolysis to generate trypsin-like activity. 329 68

Two-site methods were developed for immunoassay of acetylcholinesterase (AChE; EC 3.1.1.7) in crude extracts of rat and human tissues. A radiometric assay for human AChE utilized a specific monoclonal AChE antibody adsorbed to polystyrene microtiter wells at alkaline pH. AChE bound strongly to this antibody after 24 h at 4 degrees C. Bound enzyme was detected with an 125I-labeled antibody against a different AChE epitope. The assay signal was quasi-linearly related to AChE concentration in purified and crude samples, with a detection threshold near 100 pg. Tetrameric and dimeric AChE behaved equivalently in the assay. Two-site methods with a different pair of species-selective antibodies worked equally well for immunoassay of rat AChE. Assays of the rat enzyme showed that immunoreactivity was lost as rapidly as enzyme activity during heating to 54 degrees C. On the other hand, immunoreactivity was preserved despite loss of enzyme activity after exposure to anticholinesterases or trypsin. A biotinylated second antibody detected by alkaline-phosphatase-conjugated avidin was used to develop an AChE enzyme-linked immunosorbent assay (ELISA) with a sensitivity similar to that of the radiometric assay. Either the ELISA or the radiometric immunoassay may be useful whenever proteolysis or other mechanisms are suspected of dissociating enzyme activity and immunoreactivity. In denervated muscle and ligated peripheral nerve, application of the two-site method showed closely parallel variations in immunoreactivity and enzyme activity.
...
PMID:Two-site immunoassay for acetylcholinesterase in brain, nerve, and muscle. 329 48

Acetylcholinesterase (EC 3.1.1.7) has been shown to possess an intrinsic peptidase activity. [Chubb et al. (1983), Neuroscience 10, 1369-1383]. To examine this activity further, the breakdown of a model hexapeptide (leu-trp-met-arg-phe-ala) LWMRFA was studied. Affinity-purified eel acetylcholinesterase rapidly cleaved the hexapeptide in a trypsin-like manner to produce two peptides (LWMR and FA). Acetylcholinesterase more slowly cleaved the C-terminal alanine residue from the peptide to yield LWMRF. Although the enzyme showed preference for cleaving the hexapeptide at its C-terminal, it was also able to cleave the N-terminal leucine residue form the tryptic product LWMR. Hydrolysis of the peptide at the tryptic site (arg4-phe5) was strongly inhibited by the trypsin inhibitor diisopropylfluorophosphate. Cleavage of the C-terminal alanine was only poorly inhibited by diisopropylfluorophosphate, but more strongly inhibited by metal-ion chelating agents, and it was increased in the presence of Zn2+ and Co2+. The pH optimum for cleavage at the tryptic site was 6, while that for the carboxypeptidase site was 8-9. These results show that acetylcholinesterase can hydrolyse peptides like a trypsin-like endopeptidase and a Zn2+- or Co2+-dependent exopeptidase, and they suggest that these two peptidase activities are associated with two separate active sites on the acetylcholinesterase molecule. As both peptidase activities eluted with acetylcholinesterase from a TSK 4000SW column when it was chromatographed by high-performance liquid chromatography, it is unlikely that the presence of either peptidase activity could be attributable to a contaminant in the acetylcholinesterase preparation. We suggest that acetylcholinesterase may be involved in the breakdown of bioactive peptides or their precursors in neuroendocrine cells.
...
PMID:Acetylcholinesterase exhibits trypsin-like and metalloexopeptidase-like activity in cleaving a model peptide. 330 51

We have isolated a COOH-terminal tryptic peptide from the hydrophobic globular (5.6 S) form of Torpedo californica acetylcholinesterase that exhibits divergence in amino acid sequence from the catalytic subunit of the dimensionally asymmetric (17 S + 13 S) enzyme. The divergent peptide could be recovered from the glycophospholipid-modified 5.6 S enzyme only after treatment with phosphatidylinositol-specific phospholipase C. Upon reduction, carboxymethylation with [14C]iodoacetate, and trypsin digestion the resultant peptides were purified by gel filtration followed by high performance liquid chromatography. The high performance liquid chromatography profiles of 14C-labeled cysteine peptides from lipase-treated 5.6 S enzyme revealed unique radioactive peaks which had not been present in digests of the asymmetric form. These peaks all yielded identical amino acid sequences. The difference in chromatographic behavior of the individual peptides most likely reflects heterogeneity in post-translational processing. Gas-phase sequencing and composition analysis are consistent with the sequence: Leu-Leu-Asn-Ala-Thr-Ala-Cys. Composition includes 2-3 mol each of glucosamine and ethanolamine which is indicative of modification by glycophospholipid. Glucosamine is also present in an asparagine-linked oligosaccharide. The two forms of acetylcholinesterase diverge after the threonine residue within this peptide sequence; the hydrophobic form terminates with cysteine whereas the asymmetric form extends for 40 residues beyond the divergence. The locus of divergence and absence of any other amino acid sequence difference suggest that the molecular forms of acetylcholinesterase arise from a single gene by alternative mRNA processing.
...
PMID:Divergence in primary structure between the molecular forms of acetylcholinesterase. 333 34

A factor that stimulates the incorporation of 75Selenomethionine into the newly formed platelets of recipient mice (thrombopoietin, TPO) has been partially purified from the plasma of thrombocytopenic patients. The activity was precipitated at 60-80% ammonium sulfate saturation and further purified with hydrophobic interaction chromatography. Thrombopoietin was retained by concanavalin-A-Sepharose. Using HPLC size-exclusion chromatography, an approximate molecular weight of 40,000 dalton was calculated. The overall purification factor was about 2,100-fold. TPO was stable in a pH range from 5 to 9 and was heat-sensitive, and the biological activity was destroyed by trypsin treatment and by dithiothreitol. The partially purified molecule did not stimulate the proliferation of megakaryocyte progenitors in vitro and had no effect on the growth of erythroid or granulocyte-macrophage colonies; when administered in-vivo, TPO significantly affected the mean platelet volume and increased the number of small acetylcholinesterase cells in the bone marrow. TPO appears to be specific for the megakaryocytic lineage and active on the postmitotic compartment of megakaryocytes.
...
PMID:Partial purification and biochemical characterization of human plasma thrombopoietin. 336 51

In addition to its ability to hydrolyze acetylcholine, purified eel acetylcholinesterase possesses a trypsin-like endopeptidase activity. The tryptic activity is associated with a serine residue at a site that is distinct from the esteratic site. To label both the esteratic and tryptic sites, the enzyme was incubated with the serine hydrolase inhibitor [3H]diisopropyl fluorophosphate. This compound labelled the protein in a biphasic manner, with both slow and rapid labelling kinetics. The time course of the rapid phase was similar to the time course of inactivation of the esteratic activity. The time course of the slow phase was similar to the time course of inactivation of the tryptic activity. Labelling of the nonesteratic site was inhibited by the trypsin inhibitor N alpha-p-tosyl-L-lysine chloromethyl ketone. The total number of sites labelled by [3H]diisopropyl fluorophosphate on eel acetylcholinesterase was 2.6 mol/280,000 g protein, whereas the number of tryptic sites was less (0.52 mol/280,000 g). The results suggest that a subpopulation of acetylcholinesterase molecules may possess tryptic activity. Extensive chromatography of the purified enzyme by ion-exchange and gel filtration failed to separate the labelled tryptic component from acetylcholinesterase. On sodium dodecyl sulfate-polyacrylamide gels, the labelled tryptic component comigrated with a polypeptide of 50,000 molecular weight, which is a major proteolytic digestion product derived from the intact acetylcholinesterase monomer. Because of its localization in many noncholinergic peptide-containing cells, acetylcholinesterase could act as a neuropeptide processing enzyme in these cells.
...
PMID:Identification of a trypsin-like site associated with acetylcholinesterase by affinity labelling with [3H]diisopropyl fluorophosphate. 337 13


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---