Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Suspensions of oviduct cells were prepared by subjecting oviduct tissue to sequential incubations with EDTA, alpha-chymotrypsin, and crude collagenase, followed by a final incubation with EDTA. Cells isolated in this way incorporate mannose from exogenous GDP-mannose into mannosyl-lipid, oligosaccharide-lipid, and glycoprotein(s). Based on several criteria, the mannosyl-lipid is identical with mannosyl-phosphoryldolichol. Similarly, the oligosaccharide-lipid appears to be identical with the oligosaccharide-lipid synthesized in vitro (Lucas, J. J., Waechter, C. J., and Lennarz, W. J. (1975) J. Biol. Chem. 250, 1992-2002). In contrast, the glycoproteins are much lower in molecular weight than those labeled in cell-free preparations. Using intact oviduct cell suspensions it was found that: (a) exogenous GDP-mannose, not its breakdown products, serves as the direct mannosyl donor; (b) experiments using mixtures of known proportions of broken and intact cells, as well as studies with metabolic inhibitors, indicate that greater than 50% of the observed incorporation of mannose from GDP-mannose was catalyzed by enzymes associated with intact cells, rather than broken cells or membrane fragments; (c) incorporation of mannose from GDP-mannose into the mannosyl acceptors does not require energy and proceeds without significant uptake of GDP-mannose into trichloroacetic acid-soluble components of the cells; (d) under conditions where labeled guanosine incorporation into nucleic acids is readily detected, no incorporation of the guanosine moiety of [3H]GDP-mannose is observed. These results indicate that the enzymes catalyzing synthesis of lipid-linked intermediates involved in glycoprotein synthesis are not only associated with intracellular membranes, but with the plasma membrane as well.
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PMID:Utilization of exogenous GDP-mannose for the synthesis of mannose-containing lipids and glycoproteins by oviduct cells. 77 Apr 69

1. Single dispersed cells obtained by collagenase treatment of longitudinal muscle of rabbit small intestine were voltage clamped with low-resistance patch pipettes and membrane current was measured. 2. In cells held at -20 or -30 mV, a discharge of spontaneous transient outward currents (STOCs) was usually seen; these are believed to represent the sporadic release of calcium from storage sites in the cell in relation to TEA-sensitive, 4 AP-resistant, calcium-activated potassium channels. 3. Caffeine (20 mM) externally applied, accelerated and then abolished STOCs; carbachol (0.1 mM) had similar effects; the initial burst of STOCs was often carried on a large, temporary, outward current which could occur alone. This was suggested to be caused by the rapid release of stored calcium in relation to calcium-activated potassium channels. 4. If STOCs were abolished by caffeine (or carbachol) then carbachol (or caffeine) did not evoke outward current indicating that these drugs act on the same calcium store but by different pathways. Inclusion of ryanodine (10(-8)-10(-4) M) in the patch pipette abolished STOCs soon after establishing whole-cell recording mode; afterwards, outward current to caffeine or to carbachol could not be evoked. 5. STOCs were quickly abolished in cells patched with pipettes filled with GTP gamma S (0.1-1 mM) or Gpp(NH)p (0.1-1 mM) but were large or normal in size in cells where GDP beta S (0.1-1 mM) was included in the pipette. GTP gamma S or Gpp(NH)p in the cell abolished outward current to caffeine or to carbachol, but had no effect on calcium-activated potassium channel activity in isolated patches or on a TEA-sensitive, 4-AP-resistant, outward potassium current evoked in single cells by stepping positively from a -20 mV holding potential. These results suggest that the effect of guanine nucleotide analogues are on the calcium store rather than on calcium-activated potassium channels. 6. The effects of GTP gamma S or Gpp(NH)p could be explained if they depleted calcium stores via a G-protein mechanism; this effect may involve activation of phospholipase C enzyme (PLC) and D-myo-inositol 1,4,5-trisphosphate (IP3) production as well as a direct effect on stores. However a separate G-protein-independent pathway of activation of PLC by muscarinic receptor activation may exist as calcium release by carbachol was large or normal in cells filled with GDP beta S.
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PMID:Properties of calcium stores and transient outward currents in single smooth muscle cells of rabbit intestine. 258 96

Epidermal growth factor (EGF) regulates pancreatic acinar enzyme secretion. The mechanism of action of EGF in pancreatic acinar cells is not clear. In the present study we investigated the role of heterotrimeric GTP-binding proteins (G proteins) in EGF receptor signal transduction. Pancreatic acini were isolated from rat pancreas by collagenase digestion and permeabilized by digitonin. Activation of phosphatidylinositol 4,5-bisphosphate-specific phospholipase C (PLC) was assessed using a radioreceptor assay specific for inositol 1,4,5-trisphosphate [IP3(1,4,5)]. For measurement of amylase secretion isolated pancreatic acini were incubated with secretagogues for 30 min at 37 degrees C. Amylase released into the medium was assessed by monitoring the hydrolysis rate of p-nitrophenyl-alpha,D-maltohepatoside. The weakly hydrolyzable GTP analogue guanosine 5'-[3-O-thio]triphosphate (GTP gamma S) and guanosine 5'-diphosphate (GDP) were used to activate and inhibit G protein-mediated signal transduction, respectively. EGF (90 nM) stimulated amylase release in isolated pancreatic acini. This effect was enhanced by guanosine 5'-[3-O-thio]triphosphate (0.1 mM), which stimulates G proteins. Guanosine 5'-diphosphate (1 mM), which inhibits the activity of heterotrimeric G proteins, had no effect on basal and EGF-induced amylase release. Lower EGF concentrations (20 nM) inhibited COOH-terminal cholecystokinin octapeptide (CCK8)-induced IP3(1,4,5) production and amylase release in pancreatic acini). However, in the presence of GDP, EGF had no significant effect on CCK8-stimulated amylase release. Furthermore, coincubation of the acini with CCK8, EGF, and GDP revealed that GDP reduces the inhibitory effect of EGF on CCK8-induced IP3(1,4,5) production.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Epidermal growth factor receptor signaling in rat pancreatic acinar cells. 754 69

1. Growth hormone (GH) secretion from the anterior pituitary gland is mainly regulated by hypothalamic GH-releasing hormone (GHRH) and somatostatin (SRIF). Somatostatin reduces both spontaneous and GHRH-stimulated GH secretion. 2. Exocytosis of GH is mainly determined by the intracellular free Ca2+ concentration ([Ca2+]i), which is regulated by the influx of Ca2+ via membrane Ca2+ channels. Somatostatin reduces the influx of Ca2+ through two separate mechanisms, namely a direct action on Ca2+ channels and an indirect action on membrane potentials through the activation of K+ channels. 3. In the present experiments, somatotroph-enriched cells were obtained from the ovine pituitary gland by means of collagenase dissociation and Percoll-gradient centrifugation. Further identification was based on the effect of SRIF (10 nmol/L) on Ca2+ or K+ currents. 4. A significant reduction in Ca2+ currents and an increase in K+ currents was obtained in response to local application of SRIF (10 nmol/L), but vehicle application had no effect. The responses of Ca2+ and K+ currents to SRIF were reversible after removal of SRIF. 5. Dialysis of GTP-gamma-s (200 mumol/L) abolished the recovery phase of K+ current response to SRIF after its removal, whereas GDP-beta-s (200 mumol/L) totally blocked the response. Pretreatment of the cells with pertussis toxin (100 nmol/L) overnight abolished the Ca2+ current response to SRIF. 6. Intracellular dialysis of antibodies to alpha o, alpha i1-3, alpha i1-2 and alpha i3 subunits of the G-proteins into cells via whole-cell patch-clamp pipettes was confirmed by immunofluorescent staining of the antibodies. 7. Dialysis of anti-alpha i1-3 or anti-alpha i3 antibodies significantly attenuated the increase in the K+ current in response to 10 nmol/L SRIF, whereas neither anti-alpha o nor anti-alpha i1-2 antibodies diminished the effect of SRIF on the K+ current. 8. Dialysis of anti-alpha o antibodies significantly attenuated the reduction in the Ca2+ current that was obtained upon application of 10 nmol/L SRIF. Neither anti-alpha i1-2 nor anti-alpha i3 antibody dialysis diminished the effect of SRIF on the Ca2+ current. 9. Dialysis of the alpha o common antisense oligonucleotides (ASm) but not the alpha i3 AS significantly diminished the inhibitory effect of SRIF on the Ca2+ current. This effect of alpha o ASm dialysis occurred at 12 h incubation after dialysis, reaching a maximal level at 48 h and partially recovering at 72 h incubation. Antisense oligonucleotides specific for alpha o1 (alpha o1 AS) or alpha o2 (alpha o2 AS) were dialysed into somatotrophs and only alpha o2 AS significantly attenuated the inhibition of SRIF on the Ca2+ current. 10. It is concluded that the Gi3 protein mediates the effect of SRIF on the K+ current and that the G(o)2 protein mediates the effect of SRIF on the Ca2+ current in primary cultured ovine somatotrophs.
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PMID:G(o)2 and Gi3 proteins mediate the action of somatostatin on membrane Ca2+ and K+ currents in ovine pituitary somatotrophs. 926 41