Burn injury upregulates the activity and gene expression of the 20 S proteasome in rat skeletal muscle

2000 ◽  
Vol 99 (3) ◽  
pp. 181-187 ◽  
Author(s):  
Cheng-Hui FANG ◽  
Bing-Guo LI ◽  
David R. FISCHER ◽  
Jing Jing WANG ◽  
Herbert A. RUNNELS ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Burn Injury ◽  
Catalytic Core ◽  
Peptide Substrates ◽  
Protein Substrates ◽  
Proteolytic Pathway ◽  
Proteasome Subunits ◽  
Ubiquitin Proteasome ◽  
Dependent Protein ◽  
Increased Activity

There is evidence that burn injury stimulates ubiquitin–proteasome-dependent protein breakdown in skeletal muscle. In this proteolytic pathway, protein substrates are conjugated to multiple molecules of ubiquitin, whereafter they are recognized, unfolded and degraded by the multicatalytic 26 S protease complex. The 20 S proteasome is the catalytic core of the 26 S protease complex. The influence of burn injury on the expression and activity of the 20 S proteasome has not been reported. We tested the hypothesis that burn injury increases 20 S proteasome activity and the expression of mRNA for the 20 S proteasome subunits RC3 and RC7. Proteolytic activity of isolated 20 S proteasomes, assessed as activity against fluorogenic peptide substrates, was increased in extensor digitorum longus muscles from burned rats. Northern-blot analysis revealed that the expression of mRNA for RC3 and RC7 was increased by 100% and 80% respectively following burn injury. Increased activity and expression of the 20 S proteasome in muscles from burned rats support the concept that burn-induced muscle cachexia is at least, in part, regulated by the ubiquitin–proteasome proteolytic pathway.

Activity and expression of the 20S proteasome are increased in skeletal muscle during sepsis

1999 ◽  
Vol 277 (2) ◽  
pp. R434-R440 ◽  
Author(s):  
Scott C. Hobler ◽  
Arthur Williams ◽  
David Fischer ◽  
Jing Jing Wang ◽  
Xiaoyan Sun ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Proteolytic Activity ◽  
Blot Analysis ◽  
20S Proteasome ◽  
Catalytic Core ◽  
Proteolytic Pathway ◽  
Ubiquitin Proteasome ◽  
Dependent Protein ◽  
26S Protease

Recent studies suggest that sepsis stimulates ubiquitin-dependent protein breakdown in skeletal muscle. In this proteolytic pathway, ubiquitinated proteins are recognized, unfolded, and degraded by the multicatalytic 26S protease complex. The 20S proteasome is the catalytic core of the 26S protease complex. The role of the 20S proteasome in the regulation of sepsis-induced muscle proteolysis is not known. We tested the hypothesis that sepsis increases 20S proteasome activity and the expression of mRNA for various subunits of this complex. Proteolytic activity of isolated 20S proteasomes, assessed as activity against fluorogenic peptide substrates, was increased in extensor digitorum longus muscles from septic rats. The proteolytic activity was inhibited by specific proteasome blockers. Northern blot analysis revealed an approximately twofold increase in the relative abundance of mRNA for the 20S α-subunits RC3 and RC9 and the β-subunit RC7. However, Western blot analysis did not show any difference in RC9 protein content between sham-operated and septic rats. The increased activity and expression of the 20S proteasome in muscles from septic rats lend further support for a role of the ubiquitin-proteasome-pathway in the regulation of sepsis-induced muscle proteolysis.

Effect of guanethidine-induced adrenergic blockade on the different proteolytic systems in rat skeletal muscle

1999 ◽  
Vol 277 (5) ◽  
pp. E883-E889 ◽  
Author(s):  
Luiz Carlos C. Navegantes ◽  
Neusa M. Z. Resano ◽  
Renato H. Migliorini ◽  
Isis C. Kettelhut
Keyword(s):  
Skeletal Muscle ◽  
Adrenergic System ◽  
Initial Increase ◽  
Adrenergic Blockade ◽  
Proteolytic System ◽  
Initial Rise ◽  
Proteolytic Pathway ◽  
Increased Activity ◽  
Dependent Pathway

Overall proteolysis and the activity of skeletal muscle proteolytic systems were investigated in rats submitted to guanethidine-induced adrenergic blockade for 4 days. In soleus, overall proteolysis increased by 15–20% during the first 2 days of guanethidine treatment but decreased to levels below control values after 4 days. Extensor digitorum longus (EDL) did not show the initial increase in total proteolysis, which was already reduced after 2 days of guanethidine treatment. The initial rise in the rate of protein degradation in soleus was accompanied by an increased activity of the Ca2+-dependent proteolytic pathway. In both soleus and EDL, the reduction in overall proteolysis was paralleled by decreased activities of the Ca2+-dependent and ATP-dependent proteolytic processes. No change was observed in the activity of the lysosomal proteolytic system. Overall proteolysis in soleus and EDL from nontreated rats was partially inhibited by isoproterenol, in vitro. The data suggest an acute inhibitory control of skeletal muscle proteolysis by the adrenergic system, well evident in the oxidative muscle, with an important participation of the Ca2+-dependent pathway.

scholarly journals Evidence for an essential arginine recognition site on adenosine 3′:5′-cyclic monophosphate-dependent protein kinase of rabbit skeletal muscle

1978 ◽  
Vol 173 (2) ◽  
pp. 441-447 ◽  
Author(s):  
M Matsuo ◽  
C H Huang ◽  
L C Huang
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase ◽  
Cyclic Amp ◽  
Amino Acid Sequences ◽  
Recognition Site ◽  
Arginine Residue ◽  
Dependent Protein Kinase ◽  
Active Centre ◽  
Protein Substrates ◽  
Dependent Protein

On the basis of the chemical and structural features of the amino acid sequences in the vicinities of phosphorylatable hydroxyamino acid residues in several of the well-known protein substrates for skeletal-muscle cyclic AMP-dependent protein kinase, it is hypothesized that the phosphorylatable residue at position i and arginine residue at position i-3 of these protein substrates are located on a peptide turn on the hydrophilic protein surface. It is further hypothesized that there is an arginine-recognition site near the active centre on the protein kinase. This site is essential for the function of cyclic AMP-dependent protein kinase, for, not only does it recognize specifically the exposed arginine residue of the protein substrate, but, more importantly, via the interaction with arginine-(i′3), it may help to steer the topologically adjacent serine-i into proper orientation on the nearby active centre for phosphorylation. Model-building and kinetic data that provide support for the proposed hypotheses are presented.

Sepsis is associated with increased ubiquitinconjugating enzyme E214k mRNA in skeletal muscle

1999 ◽  
Vol 276 (2) ◽  
pp. R468-R473 ◽  
Author(s):  
Scott C. Hobler ◽  
Jing Jing Wang ◽  
Arthur B. Williams ◽  
Francesco Melandri ◽  
Xiaoyan Sun ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Cecal Ligation ◽  
Sham Operation ◽  
Protein Breakdown ◽  
Protein Levels ◽  
Proteolytic Pathway ◽  
Ubiquitin Conjugating Enzyme ◽  
Slow Twitch Muscle ◽  
Dependent Protein ◽  
Slow Twitch

Previous studies provided evidence that sepsis is associated with increased ubiquitin-proteasome-dependent protein breakdown in skeletal muscle. The 14-kDa ubiquitin-conjugating enzyme (E214k) has been proposed to be a key regulator of the ubiquitin proteolytic pathway. We tested the hypothesis that E214k message and protein levels are increased in skeletal muscle during sepsis. Sepsis was induced in rats by cecal ligation and puncture (CLP). Control rats were sham operated. E214k mRNA and protein levels were quantitated after Northern and Western blot analysis, respectively, 16 h after CLP or sham operation. Sepsis resulted in a 70% increase in the 1.2-kb E214k transcript in the fast-twitch extensor digitorum longus muscle, whereas no changes were seen in the slow-twitch soleus muscle. E214k protein levels were not influenced by sepsis in any of the muscles studied. Although the changes in the expression of the E214k 1.2-kb transcript paralleled the differential effect of sepsis on protein breakdown in fast- and slow-twitch muscle, the potential role of E214k in the regulation of sepsis-induced muscle proteolysis needs to be interpreted with caution, because the results demonstrated that increased message levels were not associated with increased E214kprotein levels.

scholarly journals Influence of a mutation in the ATP-binding region of Ca2+/calmodulin-dependent protein kinase II on its interaction with peptide substrates

2004 ◽  
Vol 378 (2) ◽  
pp. 391-397 ◽  
Author(s):  
Mullasseril PRASEEDA ◽  
Kurup K. PRADEEP ◽  
Ananth KRUPA ◽  
S. Sri KRISHNA ◽  
Suseela LEENA ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Catalytic Domain ◽  
3D Structure ◽  
Atp Binding ◽  
Binding Region ◽  
Dependent Protein Kinase ◽  
Catalytic Core ◽  
Peptide Substrates ◽  
Protein Kinase Ii ◽  
Dependent Protein

CaMKII (Ca2+/calmodulin-dependent protein kinase II) is expressed in high concentrations in the brain and is found enriched in the postsynaptic densities. The enzyme is activated by the binding of calmodulin to the autoregulatory domain in the presence of high levels of intracellular Ca2+, which causes removal of auto-inhibition from the N-terminal catalytic domain. Knowledge of the 3D (three-dimensional) structure of this enzyme at atomic resolution is restricted to the association domain, a region at the extreme C-terminus. The catalytic domain of CaMKII shares high sequence similarity with CaMKI. The 3D structure of the catalytic core of CaMKI comprises ATP- and substrate-binding regions in a cleft between two distinct lobes, similar to the structures of all protein kinases solved to date. Mutation of Glu-60, a residue in the ATP-binding region of CaMKII, to glycine exerts different effects on phosphorylation of two peptide substrates, syntide and NR2B (N-methyl-d-aspartate receptor subunit 2B) 17-mer. Although the mutation caused increases in the Km values for phosphorylation for both the peptide substrates, the effect on the kcat values for each was different. The kcat value decreased in the case of syntide, whereas it increased in the case of the NR2B peptide as a result of the mutation. This resulted in a significant decrease in the apparent kcat/Km value for syntide, but the change was minimal for the NR2B peptide. These results indicate that different catalytic mechanisms are employed by the kinase for the two peptides. Molecular modelling suggests structural changes are likely to occur at the peptide-binding pocket in the active state of the enzyme as a consequence of the Glu-60→Gly mutation.

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