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

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

The structural features of "natural"human monocyte-mediated cytotoxicity towards K-562 leukemia cells in vitro have been examined. Human monocytes bound K-562 cells firmly and mediated a slow cytolysis of the leukemia cells. Monocyte binding of target cells was shown to be trypsin sensitive. Freeze-fracture and thin section electron microscopy revealed that effector and target cells were separated by a irregular space larger than 20 nm. There was no evidence for the involvement of specialized membrane junctions or organelle transfer between monocytes and target cells. Scanning electron microscopy revealed that K-562 cells bound to monocytes progressively lost their microvilli. This process started in the membrane areas close to the effector-target cell interaction. The results suggest that binding of target cells by monocytes followed by action of short-range soluble cytotoxic mediators may be the mechanism for the monocyte-mediated cytotoxicity towards K-562 cells.
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PMID:Human monocyte mediated cytotoxicity to K-562 cells: a freeze-fracture-, scanning- and thin section electron microscopic study. 720 21

The major intrinsic protein from spinach chloroplast membranes, the light-harvesting chlorophyll a/b-protein complex, contains two distinct polypeptides of Mr 23,500 and 26,000 and 31% lipid by weight, comprising five diacyl lipids and seven chlorophylls, together with some carotenoids, per 26,000-Mr polypeptide. The chlorophyll a/b ratio is 1.1. Low-temperature fluorescence emission spectra of the light-harvesting complex revealed a major peak at 681 nm with a shoulder of variable intensity at 695 nm. The 695-nm emission has been correlated with a progressive aggregation of the complex into two-dimensional, semi-crystalline sheets. To determine the role of the light-harvesting complex in cation-dependent thylakoid stacking, the purified complex has been quantitatively incorporated into liposomes containing the four major chloroplast diacyl lipids using a simple freeze-thaw technique. The proteoliposomes appeared largely as unilamellar vesicles, with diameters between 0.1 and 0.8 micron. Freeze-fracture analysis showed intramembrane particles of 8-10 nm corresponding to the incorporated complex. Both monovalent and divalent cations caused an immediate aggregation of the proteoliposomes, which was reversed at low cation concentrations and was largely inhibited by prior trypsin treatment. Since lipid vesicles themselves showed none of these effects, it is concluded that surface-exposed polypeptide regions of the light-harvesting complex are directly involved in thylakoid stacking in vivo.
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PMID:The role of the light-harvesting chlorophyll a/b protein complex in chloroplast membrane stacking. Cation-induced aggregation of reconstituted proteoliposomes. 739 45

Several subpopulations of cells were isolated from trypsin-dissociated embryonic (14 days) chick retinas. The cells of each subpopulation differed in associative behavior measured by cell aggregation and stationary culture assays and in glycoproteins that contain glucosamine. Freeze-fracture analysis showed that these populations also differed in intramembrane particle content.
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PMID:Cells isolated from the embryonic neural retina differ in behavior in vitro and membrane structure. 740 67

Fc and C3 receptors, which are characteristically present on macrophages, could not be demonstrated on osteoclasts maintained in situ on their normal substrates when assayed for by use of sheep red blood cells coated with immunoglobulin (Shapiro et al. 1979). The present study tested the hypotheses that Fc receptors are present only on the osteoclast surface adjacent to bone and that Fc receptors on osteoclasts can be uncovered by enzymes or stimulated to appear. Freeze-dried, inverted osteoclasts (and osteoblasts) obtained from the endocranium of newborn rats were tested for Fc receptors using the rosette assay and examined by scanning electron microscopy. No rosettes were observed on the surfaces of the osteoclasts that had been approximal to the bone. Bone specimens were cultured for 30 min at 37 degrees C in control medium, or in medium with the addition of 10, 50 or 100 microgram/ml trypsin, 0.5 U/ml parathyroid extract (PTE), or 0.5 or IU/ml parathyroid hormone 1--34 (PTH). Additionally, two week-old rats were injected intraperitoneally with PTE (1.5 U/g body weight or 1 USP/g body weight) or with PTH (1 U/g body weight) or with vehicle alone, 6 h before sacrifice. The specimens were assayed for Fc receptors and examined by scanning electron microscopy. Macrophages were always used as controls for the assay. No rosettes were present on osteoclasts subjected to any of these treatments. Accordingly, the hypotheses were not supported.
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PMID:Lack of Fc receptors on osteoclasts. 745 90

Cockerels from lines of White Plymouth Rock chickens selected for 33 generations for high (HW) or low (LW) 8-wk BW were used in this experiment. Either Diet A (a diet similar to that under which selection had been conducted) or Diet B (containing 20% more CP and 20% more ME) were consumed ad libitum for the entire experiment (hatch to 6 wk of age). Body weights, organ weights, and enzymes present in the gastrointestinal tract (GIT) contents and pancreas were measured. There were differences between lines and between diets for BW and relative breast weight. Several organs (heart, lung, liver, and pancreas) generally remained a constant proportion of BW in both lines. Relative weight of the GIT was greater in HW cockerels from hatch to 10 d of age, after which relative GIT weight was greater in LW cockerels. Relative weight of the GIT was also affected after 10 d of age by density of feed. Interactions between line and diet were present for many digestive enzyme measurements, necessitating analyses within each main effect. All pancreatic enzymes except relative trypsin were higher in HW than LW cockerels. Although small intestine enzyme activities were higher in Line HW than LW, Cockerels on a relative basis the difference was present only for trypsin activity. Cockerels fed Diet B had higher levels of pancreatic chymotrypsin activity and of total and relative lipase activities than those fed Diet A.
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PMID:Enzyme activity and organ development in newly hatched chicks selected for high or low eight-week body weight. 760 51

The amino acid sequence of copper pheasant lysozyme was analyzed. Carboxymethylated lysozyme was digested with trypsin and the resulting peptides were sequenced. The established amino acid sequence had three amino acid substitutions at positions 20, 77, and 113 for Lady Amherst's pheasant lysozyme and seven amino acid substitutions at positions 3, 15, 20, 41, 113, 121, and 124 for hen lysozyme. Phenylalanine at position 20 was newly detected in avian lysozyme.
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PMID:The amino acid sequence of copper pheasant lysozyme. 776 70

The formation of transgenic chimeric chickens for use in developmental studies and as intermediates in the production of transgenic chickens requires the incorporation of stably transfected blastodermal cells into a chimera. To obtain blastodermal cells, area pellucidae of stage X (Eyal-Giladi and Kochav, Dev. Biol. 49:321-337, 1976:E.-G.&K.) embryos were collected from unincubated, freshly oviposited Barred Plymouth Rock eggs and dissociated in 0.25% trypsin/0.04% EDTA (w/v) and 2% (v/v) chicken serum in phosphate-buffered saline (Ca2+ and Mg2+ free) at 4 degrees C for 10 min. The blastodermal cells were suspended in Dulbecco's Modified Eagle's Medium (DMEM) and transfected by lipofection with superhelical pmiwZ, a plasmid containing a hybrid lacZ gene encoding bacterial beta-galactosidase (beta-gal) under the control of a chicken beta-actin/Rous sarcoma virus promoter. A mixture of 2.5 micrograms Lipofectin and 1.56 micrograms pmiwZ in 250 microliters DMEM was incubated for 30 min at 37 degrees C and added to 500 microliters of 20-40,000 cells in suspension. Cells incubated with the transfection reagents in the presence or absence of pmiwZ were either plated and cultured for 48 h at 37 degrees C in 5% CO2/95% air, or injected through a shell window into the subgerminal cavity of White Leghorn stage X (E.-G.&K.) embryos previously exposed to 500-600 rads from a 60Co source, after which the window was sealed and the egg incubated at 38 +/- 1 degrees C for 72 h.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Efficient incorporation of transfected blastodermal cells into chimeric chicken embryos. 829 32

The vaccinia virus forms two morphologically distinct infectious virus particles: intracellular mature virus (IMV) and extracellular enveloped virus (EEV). The envelope of EEV is a Golgi-derived membrane (wrapping membrane). A mutant (vRB10) lacks the ability to form the EEV. In medium containing a neutralizing antibody (2D5mAb), the vRB10 mutant was diluted out from infected cells, whereas the IHD-J strain of vaccinia virus replicated well. The result indicated that the 2D5mAb specifically neutralized the IMV. The 2D5-resistant EEV appeared at 6-7 hr postinfection, and over 65% of infectious virus in the culture fluid was EEV at 48 hr after infection. The EEV was resistant not only to the 2D5mAb but also against several neutralizing antibodies, including polyclonal antivaccinia serum reactive with proteins of the wrapping membrane. Freeze-thawing and other procedures that may damage the wrapping membrane converted the EEV to a form susceptible to the antibodies. Since specific infectivity was not affected by the damage or by exposure to antibody against the wrapping membrane proteins, the wrapping membrane did not directly participate in penetration. The infection process of vaccinia virus was analyzed by comparison of responses to acid treatment between normal IMV and trypsin-treated IMV. Proteolytically activated IMV infected rapidly responding to acid. The protected form virus, which was noninfectious under usual conditions, was proteolytically activated on cell membrane then responded to the acid. Proteolysis activated the virus, and an acidic condition accelerated fusion between the activated IMV and plasma membrane. The virus in the EEV wrapping membrane was the activated form that has the capacity to fuse with the cell membrane. However, the infection of intact EEV was more sensitive against lysosomotropic agents (NH4Cl, neutral red) than that of the trypsin-activated IMV. Resistance to the 2D5mAb, sensitivity to lysosomotropic agents, and acceleration of infection by acid suggested that the intact EEV penetrated by virus-endosome membrane fusion. The combined effect of the presence of wrapping membrane and the process of internalization via an endocytic mechanism rendered EEV resistant to neutralizing antibodies.
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PMID:Extracellular enveloped vaccinia virus escapes neutralization. 861 Apr 39

Trypsin and chymotrypsin have specificity pockets of essentially the same geometry, yet trypsin is specific for basic while chymotrypsin for bulky hydrophobic residues at the P1 site of the substrate. A model by Steitz, Henderson and Blow suggested the presence of a negative charge at site 189 as the major specificity determinant: Asp189 results in tryptic, while the lack of it chymotryptic specificity. However, recent mutagenesis studies have shown that a successful conversion of the specificity of trypsin to that of chymotrypsin requires the substitution of amino acids at sites 138, 172 and at thirteen other positions in two surface loops, that do not directly contact the substrate. For further testing the significance of these sites in substrate discrimination in trypsin and chymotrypsin, we tried to change the chymotrypsin specificity to trypsin-like specificity by introducing reverse substitutions in rat chymotrypsin. We report here that the specificity conversion is poor: the Ser189Asp mutation reduced the activity but the specificity remained chymotrypsin-like; on further substitutions the activity decreased further on both tryptic and chymotryptic substrates and the specificity was lost or became slightly trypsin-like. Our results indicate that in addition to structural elements already studied, further (chymotrypsin) specific sites have to be mutated to accomplish a chymotrypsin --> trypsin specificity conversion.
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PMID:Attempts to convert chymotrypsin to trypsin. 863 80

Trypsin and chymotrypsin have specificity pockets of essentially the same geometry, yet trypsin is specific for basic while chymotrypsin for bulky hydrophobic residues at the P1 site of the substrate. A model by Steitz, Henderson and Blow suggested the presence of a negative charge at site 189 as the major specificity determinant: Asp189 results in tryptic, while the lack of it chymotryptic specificity. However, recent mutagenesis studies have shown that a successful conversion of the specificity of trypsin to that of chymotrypsin requires the substitution of amino acids at sites 138, 172 and at thirteen other positions in two surface loops, that do not directly contact the substrate. For further testing the significance of these sites in substrate discrimination in trypsin and chymotrypsin, we tried to change the chymotrypsin specificity to trypsin-like specificity by introducing reverse substitutions in rat chymotrypsin. We report here that the specificity conversion is poor: the Ser189Asp mutation reduced the activity but the specificity remained chymotrypsin-like; on further substitutions the activity decreased further on both tryptic and chymotryptic substrates and the specificity was lost or became slightly trypsin-like. Our results indicate that in addition to structural elements already studied, further (chymotrypsin) specific sites have to be mutated to accomplish a chymotrypsin-->trypsin specificity conversion.
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PMID:Attempts to convert chymotrypsin to trypsin. 861 81


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