Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.27 (thermolysin)
 1,894 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Myosin subfragment-1 from rabbit skeletal muscle was digested by thermolysin at 25 degrees, 12 degrees and 0 degree C. Thermolysin cleaves subfragment-1 heavy chain into two stable fragments, 28 kDa and 70 kDa, aligned in this order from the N-terminus [Applegate, D. & Reisler, E. (1983) Proc. Natl Acad. Sci. USA 80, 7109-7112]. The rate of digestion at 25 degrees C was significantly increased in the presence of MgATP and somewhat less in the presence of MgADP, or magnesium pyrophosphate. This activating effect of the nucleotides was decreased at 12 degrees C and completely eliminated at 0 degrees C. The results can be explained by assuming that there are two subfragment-1 conformers [Shriver, J. W. & Sykes, B. D. (1981) Biochemistry 20, 2004-2012], and that both the addition of ATP or its analogs, and lowering the temperature, shift the conformational equilibrium in the direction that is more susceptible to thermolysin. Actin inhibited thermolysin digestion of subfragment-1 at all three temperatures studied. Actin inhibition can be explained either by shifting the equilibrium of the conformers in the direction of the less susceptible form or by direct interference of actin with the binding of thermolysin to subfragment-1. Actin inhibition of thermolysin digestion also prevailed when subfragment-1 was in a ternary complex with nucleotide and actin, in both the strongly and weakly attached states. Similarly, actin inhibited the digestion of subfragment-1 modified by 4-phenylenedimaleimide [corrected], which also forms a weakly attached complex with actin. No difference could be found in the accessibility of the thermolysin-susceptible site of subfragment-1 at the 28-70 kDa junction in either rigor, strongly or weakly attached states, which indicates the similarity of the structure proximal to this specific site in the three attached states.
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PMID:Effect of nucleotides, actin and temperature on thermolysin digestion of myosin subfragment-1. 220 15

Rat fat cells contain three species of spontaneously active inhibitor proteins of protein phosphatase 1, as resolved by SDS-PAGE, with apparent molecular masses of 40 kDa, and 28 kDa respectively. The 33-kDa, thermostable inhibitor was highly purified from bovine adipose tissue and shown to be very similar to inhibitor-2 of skeletal muscle. It was phosphorylated, on threonine only, by glycogen synthase kinase 3. It formed an inactivated complex with protein phosphatase 1, that was reactivated by incubation with ATP-Mg and glycogen synthase kinase 3. By gel filtration it had a Stokes radius of 3.4 nm. Peptide and phosphopeptide maps, generated by Staphylococcus aureus V8 proteinase, trypsin or thermolysin, of the inhibitor and of the skeletal muscle inhibitor-2 were similar. The 40-kDa inhibitor, which was denatured by boiling, represents a novel protein phosphatase inhibitor protein or an undegraded precursor of inhibitor-2. The total activity of inhibitor-2-like material (thermostable and macromolecular) in an adipocyte cytosol extract corresponded to an intracellular concentration of 0.3 microM inhibitor-2.
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PMID:Adipose tissue protein phosphatase inhibitor-2. 282 48

Kallikrein-4 (KLK4) is a serine proteinase believed to be important in the normal development of dental enamel. We isolated native KLK4 from developing pig enamel and expressed four recombinant forms. Pig KLK4 was expressed in bacteria with and without the propeptide, and in two eukaryotic systems. Recombinant pig KLK4 was secreted as a zymogen by '293' cells and purified. The proKLK4 was activated in vitro by thermolysin and recombinant pig enamelysin, but not by native KLK4. These results were confirmed using a fluorescent peptide analog of the KLK4 propeptide-enzyme junction. Native KLK4 appears as a doublet at 37 kDa and 34 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Removal of N-linked oligosaccharides by digestion with deglycosidase-F reduced the doublet to a single band at approximately 28 kDa, demonstrating that the active enzyme is glycosylated, and that the 37 kDa and 34 kDa forms differ only in their number of glycosylations. Deglycosylation was also associated with a loss of proteolytic activity. We digested recombinant pig amelogenin with native KLK4 and characterized the cleavage products by N-terminal sequencing and mass spectrometry. Eleven cleavage sites in the amelogenin protein were identified, demonstrating that KLK4 degrades amelogenin and is likely to participate in the degradation of enamel proteins in vivo.
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PMID:Porcine kallikrein-4 activation, glycosylation, activity, and expression in prokaryotic and eukaryotic hosts. 1266 66