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)

1. Hydrolysis of the myosins from smooth and from skeletal muscle by a rat trypsin-like serine proteinase and by bovine trypsin at pH 7 is compared. 2. Proteolysis of the heavy chains of both myosins by the rat enzyme proceeds at rates approx. 20 times faster than those obtained with bovine trypsin. Whereas cleavage of skeletal-muscle myosin heavy chain by both enzymes results in the generation of conventional products i.e. heavy meromyosin and light meromyosin, the heavy chain of smooth-muscle myosin is degraded into a fragment of mol. wt. 150000. This is dissimilar from heavy meromyosin and cannot be converted into heavy meromyosin. It is shown that proteolysis of the heavy chain takes place in the head region. 3. The 'regulatory' light chain (20kDa) of smooth-muscle myosin is degraded very rapidly by the rat proteinase. 4. The ability of smooth-muscle myosin to have its ATPase activity activated by actin in the presence of a crude tropomyosin fraction on introduction of Ca2+ is diminished progressively during exposure to the rat proteinase. The rate of loss of the Ca2+-activated actomyosin ATPase activity is very similar to the rate observed for proteolysis of the heavy chain and 3-4 times slower than the rate of removal of the so-called 'regulatory' light chain. 5. The significance of these findings in terms of the functional organization of the smooth muscle myosin molecule is discussed. 6. Since the degraded myosin obtained after exposure to very small amounts of the rat proteinase is no longer able to respond to Ca2+, i.e. the functional activity of the molecule has been removed, the implications of a similar type of proteolysis operating in vivo are considered for myofibrillar protein turnover in general, but particularly with regard to the initiation of myosin degradation, which is known to take place outside the lysosome (i.e. at neutral pH).
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PMID:Degradation of smooth-muscle myosin by trypsin-like serine proteinases. 612 14

Contractile and regulatory proteins were prepared from bovine aortic intima, and actin from bovine stomach smooth and rabbit skeletal muscles. In the desensitized and reconstituted actomyosin system, the superprecipitation activity was measured by the turbidity method. Superprecipitation of each system was not exhibited even in the presence of Ca ions, but was observable only in the presence of tropomyosin and Ca ions, while 20,000-dalton light chain of myosin remained dephosphorylated during the reaction. Addition of tropomyosin to the reconstituted acto-myosin digest system (trypsin-digested myosin was devoid of 20,000-dalton light chain) also restored the Ca2+-sensitivity. These results indicate that the phosphorylation of myosin light chain is not a crucial step in the contraction of aortic intima smooth muscle. For full activation of the actin-myosin-ATP interaction, additional factors other than the myosin light chain kinase are required, although some contribution of the kinase to the full activation cannot be ruled out.
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PMID:Ca2+ regulation not associated with phosphorylation of myosin light chain in aortic intima smooth muscle. II. Effects of regulatory proteins on the actin-myosin interaction. 622 81

The steady-state level and synthesis of a pair of polypeptides of Mr 33,000 and 35,000 in chicken embryo fibroblasts (CEF) transformed by Rous sarcoma virus (RSV) are significantly decreased relative to normal CEF; however, the decrease is more pronounced in the case of the Mr 35,000 polypeptide. These polypeptides have been identified as the alpha and beta subunits of CEF tropomyosin by selective staining with tropomyosin antibody, two-dimensional gel electrophoresis, partial peptide analysis, and solubility properties. The decrease in tropomyosin is shown to be a transformation-specific phenomenon in that it does not occur after infection with a virus deleted in src sequences. Decreased synthesis of tropomyosin is also observed in quail cells transformed by MC29 (a retrovirus with a different onc gene than that in RSV) and also in chemically transformed quail cells. The decreased in tropomyosin is probably not a direct result of the disruption of the microfilament system in transformed cells because disruption of the microfilament system with trypsin or cytochalasin B in normal CEF does not lead to a decrease in tropomyosin synthesis. A decrease in tropomyosin in CEF after transformation may be a result of a pleiotropic effect that results in the transcriptional inactivation not only of the tropomyosin gene but also of the fibronectin and procollagen genes described by others.
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PMID:Tropomyosin is decreased in transformed cells. 627 10

Glycerinated muscle fiber from rabbit psoas muscle often lost Ca2+-dependent regulation of its contraction with long-term extraction in a 50% glycerol solution containing 5 mM EGTA at -20 degrees C, designated as Ca2+-insensitive muscle fiber (CaIS-fiber). About 30 or 40% of glycerinated muscle fibers were CaIS-fibers after 1 to 3 months in the glycerol solution. We investigated the cause of the loss of Ca2+-sensitivity of the glycerinated muscle fiber by tension mechanogram and SDS polyacrylamide gel electrophoresis. This natural CaIS-fiber showed a new band of 30K daltons peptide on SDS gels. On the other hand, Ca2+-sensitive fiber (CaS-fiber) changed to CaIS-fiber by trypsin digestion for 40 sec. The tryptic CaIS-fiber had no troponin C and 30K daltons peptide bands in the electrophoretograms. Incubating with 2 mM CaCl2 for 40 hr at 25 degrees C, CaS-fiber changed easily to CaIS-fiber which had 30K daltons peptide and faint troponin T and I bands, as in natural CaIS-fiber. All CaIS-fibers could recover their Ca2+-dependent regulation by incubating with native tropomyosin from rabbit skeletal muscle for 2 days at 4 degrees C. These results indicate that the loss of Ca2+-dependent regulation of glycerinated muscle fiber is due to degradation of regulatory protein system by endogenous Ca2+-activated proteolytic enzymes.
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PMID:Loss of Ca2+-dependent regulation in glycerinated skeletal muscle contraction. 635 84

Fragments of rabbit skeletal muscle Ca2+-binding subunit of troponin (TnC), obtained by cleavage with trypsin, thrombin, and CNBr, were tested for their ability to form binary and ternary complexes with ATPase inhibitory subunit (TnI) and tropomyosin-binding subunit (TnT) and their ability to replace TnC in reversing TnI inhibition of actomyosin ATPase activity. Three regions of TnC were found to be involved in interaction with TnI. Regions near Ca2+-binding sites II and III require Ca2+ for the interaction, while a third region near Ca2+-binding site IV binds TnI whether or not Ca2+ is present. The TnT binding site has been localized in the NH2-terminal half of TnC. Several of the TnC fragments form soluble ternary complexes with TnI and TnT. Fragments that contain amino acid residues 89-100 and at least one pair of Ca2+-binding sites are able to reverse the TnI inhibition of actomyosin ATPase activity, which exhibits the same [Ca2+]1/2 regardless of which of the Ca2+-binding sites are present in the fragment.
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PMID:Proteolytic fragments of troponin C. Interactions with the other troponin subunits and biological activity. 645 9

Structural changes in tropomyosin from rabbit skeletal muscle were studied by the tryptic digestion method, which is an application of the quantitative enzyme-probe method recently developed by Ueno and Harrington [Ueno, H., & Harrington, W.F. (1984) J. Mol. Biol. 173, 35-61]. Effects of ionic strength, temperature, and an interchain disulfide bond at Cys-190 on the structure of tropomyosin were examined. A region of high susceptibility to trypsin was found to be localized in the middle portion of the molecule, and its susceptibility increased on lowering ionic strength and/or raising temperature. With the introduction of a disulfide bond at Cys-190, cleavage on the N-terminal side of Cys-190 was accelerated. The results suggest that skeletal muscle tropomyosin is flexible in the middle of the molecule in contrast to the flanking N- and C-terminal trypsin-resistant segments.
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PMID:Local structural changes in tropomyosin detected by a trypsin-probe method. 649 61

Isolated smooth muscle cells and cell fragments prepared by glycerination and subsequent homogenization will contract to one-third their normal length, provided Ca++ and ATP are present. Ca++-independent contraction was obtained by preincubation in Ca++ and ATP gamma S, or by addition of trypsin-treated myosin light chain kinase (MLCK) that no longer requires Ca++ for activation. In the absence of Ca++, myosin was rapidly lost from the cells upon addition of ATP. Glycerol-urea-PAGE gels showed that none of this myosin is phosphorylated. The extent of myosin loss was ATP- and pH-dependent and occurred under conditions similar to those previously reported for the in vitro disassembly of gizzard myosin filaments. Ca++-dependent contraction was restored to extracted cells by addition of gizzard myosin under rigor conditions (i.e., no ATP), followed by addition of MLCK, calmodulin, Ca++, and ATP. Function could also be restored by adding all these proteins in relaxing conditions (i.e., in EGTA and ATP) and then initiating contraction by Ca++ addition. Incubation with skeletal myosin will restore contraction, but this was not Ca++-dependent unless the cells were first incubated in troponin and tropomyosin. These results strengthen the idea that contraction in glycerinated cells and presumably also in intact cells is primarily thick filament regulated via MLCK, that the myosin filaments are unstable in relaxing conditions, and that the spatial information required for cell length change is present in the thin filament-intermediate filament organization.
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PMID:Regulation of contraction and thick filament assembly-disassembly in glycerinated vertebrate smooth muscle cells. 668 23

Myofibrils that had been exposed to rat pancreatic trypsin-like serine proteinase and to beef heart Ca2+-activated thiol proteinase were examined in the electron microscope and by SDS-gel electrophoresis. The former enzyme caused more extensive disruption of the ultrastructure and degraded more of the myofibril proteins than the CA2+-activated proteinase. The susceptibilities of individual purified proteins to the two enzymes were also compared. Myosin was virtually resistant to the Ca2+-activated enzyme but with smooth muscle myosin/rat serine proteinase at a ratio of 20000/1, heavy chain degradation took place very rapidly and the ability of the degraded myosin to have its ATPase activity activated by actin in the presence of Ca2+ was lost at a similar rate. G-actin, troponins T and I and alpha-actinin were also degraded readily by the trypsin-like proteinase whereas tropomyosin, a negatively charged rodlike protein, was more resistant. The cellular location of both proteinases remains to be established but from these results obtained in vitro, consideration is given to whether these types of proteinase might work cooperatively in vivo to bring about the disassembly and turnover of myofibrillar proteins that is known to take place outside the lysosomes in muscle.
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PMID:Proteolysis of myofibrillar proteins at neutral pH. 704 97

The effects of the Ca2+-activated cysteine proteinase, the rat trypsin-like serine proteinase and bovine trypsin on myofibrillar proteins from rabbit skeletal muscle are compared. 2. Myofibrils that had been treated at neutral pH with the Ca2+-dependent proteinase and with the rat enzyme were (a) analyzed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and (b) examined in the electron microscope. Treatment with each proteinase resulted in the loss of the Z-discs, but the rat enzyme caused much more extensive disruption of the ultrastructure and degraded more of the myofibrillar proteins. 3. Purified F-actin was almost totally resistant to the proteinases, whereas G-actin was degraded by the rat trypsin-like proteinase at a rate approx. 15 times faster than was obtained with bovine trypsin. 4. Similar results were obtained with alpha-actinin, whereas tropomyosin was degraded more readily by bovine trypsin than by the rat trypsin-like proteinase. 5. The implications of these findings for the non-lysosomal breakdown of myofibrillar proteins in vivo are considered.
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PMID:Degradation of myofibrillar proteins by trypsin-like serine proteinases. 704 73

Endometrial proteins showing cyclic expression during the normal menstrual cycle were localized on two-dimensional (2-D) electrophoresis gels separating proteins with isoelectric points (pl) ranging from 3.5 to 7 and relative molecular weights ranging from 10 to 300 kDa. Menstrual cycle-related proteins were excised from several 2-D gels, concentrated by one-dimensional (1-D) sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis, and cleaved in situ by trypsin. The tryptic fragments were extracted and separated by reverse phase high performance liquid chromatography (RP-HPLC). Finally, the partial amino-terminal amino acid sequence of selected tryptic fragments were determined for each protein. We aimed at characterizing the 21 menstrual cycle-related proteins that were visible on silver-stained 2-D electrophoresis gels. Of the proteins being maximally synthesized in the proliferative phase endometrium, we identified proteins associated mainly with the cytoskeleton: vimentins, keratin, tropomyosin and tubulin, but also proteins such as proliferating cell nuclear antigen and beta-galactoside binding lectin. The partial amino acid sequences for another two proteins did not match any protein sequence in the Protein Identification Resource (PIR) and Swissprot databases. In the group of proteins having maximal synthesis in the secretory phase endometrium, we identified creatine kinase chain B and an isocitrate dehydrogenase-homologous protein, both of which are involved in energy metabolism. However, we also identified the annexin IV precursor, the 14-3-3 protein homologue also called stratifin or the epithelial cell marker protein 1 and the 21K tumour protein. Finally, four of the proteins were present in too low amounts to allow characterization. Interestingly, most of the identified proteins have not previously been described as having a menstrual cycle-related synthesis in the human endometrium. It may be considered that the concentration of some of the cycle-related proteins may be used in clinical situations to reflect specific endometrial phases.
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PMID:Human endometrial proteins with cyclic changes in the expression during the normal menstrual cycle: characterization by protein sequence analysis. 856 10


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