• Journal of Cancer Prevention
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• v.23 no.4
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• pp.153-161
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• 2018

Imbalance of protein homeostasis (proteostasis) is known to cause cellular malfunction, cell death, and diseases. Elaborate regulation of protein synthesis and degradation is one of the important processes in maintaining normal cellular functions. Protein degradation pathways in eukaryotes are largely divided into proteasome-mediated degradation and lysosome-mediated degradation. Proteasome is a multisubunit complex that selectively degrades 80% to 90% of cellular proteins. Proteasome-mediated degradation can be divided into 26S proteasome (20S proteasome + 19S regulatory particle) and free 20S proteasome degradation. In 1980, it was discovered that during ubiquitination process, wherein ubiquitin binds to a substrate protein in an ATP-dependent manner, ubiquitin acts as a degrading signal to degrade the substrate protein via proteasome. Conversely, 20S proteasome degrades the substrate protein without using ATP or ubiquitin because it recognizes the oxidized and structurally modified hydrophobic patch of the substrate protein. To date, most studies have focused on protein degradation via 26S proteasome. This review describes the 26S/20S proteasomal pathway of protein degradation and discusses the potential of proteasome as therapeutic targets for cancer treatment as well as against diseases caused by abnormalities in the proteolytic system.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.1
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• pp.133-140
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• 2005

Although amino acids are substrates for the synthesis of proteins and nitrogen-containing compounds, it has become more and more clear over the years that these nutrients are also extremely important as regulators of body protein turnover. The branched-chain amino acids (BCAAs) together or simply leucine alone stimulate protein synthesis and inhibit protein breakdown in skeletal muscle. However, it was only recently that the mechanism(s) involved in the regulation of protein turnover by BCAAs has begun to be defined. The acceleration of protein synthesis by these amino acids seems to occur at the level of peptide chain initiation. Oral administration of leucine to food-deprived rats enhances muscle protein synthesis, in part, through activation of the mRNA binding step of translation initiation. Despite our knowledge of the induction of protein synthesis by BCAAs, there are few studies on the suppression of protein degradation. The recent findings that oral administration of leucine rapidly reduced $N^{\tau}$-methylhistidine (3-methylhistidine; MeHis) release from isolated muscle, an index of myofibrillar protein degradation, indicate that leucine suppresses myofiblilar protein degradation. The details of the molecular mechanism by which leucine inhibits proteolysis is just beginning to be elucidated. The purpose of this report was to review the current understanding of how BCAAs act as regulators of protein turnover.

• Journal of the Korean Magnetic Resonance Society
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• v.26 no.4
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• pp.66-70
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• 2022

Protein structure determination using NMR spectroscopy requires a suite of heteronuclear 3-D NMR experiments that can take a couple of weeks for completion. During the experiments, protein samples may suffer from slow degradation due to co-purifying proteases, which complicates and slows down the assignment procedure. Here we describe a practical protocol to avoid unwanted proteolysis during the experiment.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.5
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• pp.616-622
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• 2011

This study evaluated the fermentation characteristics and protein degradation dynamics of wet green tea grounds (WGTG) silage. The WGTG was ensiled with distilled water (control), or lactic acid bacteria (LAB), enzyme (E), formic acid (FA) and formaldehyde (FD) prior to ensiling. Three bag silos for each treatment were randomly opened at 0, 3, 7, 14, 28 and 60 days after anaerobic storage. For all the treatments, except for FA, there was a rapid decline in pH during the first 7 days of ensiling. LAB treatment had higher lactic acid content, lower ammonia-N ($NH_3$-N) and free-amino nitrogen (FAA-N) contents than other treatments (p<0.05). E treatment had higher lactic acid, water-soluble carbohydrates (WSC) and non-protein nitrogen (NPN) content than the control (p<0.05). FA treatment had higher $NH_3$-N and FAA-N content than the control (p<0.05). FD treatment had lower NPN and FAA-N content than the control, but it did not significantly inhibit the protein degradation when compared to LAB treatment (p>0.05). Results indicate that LAB treatment had the best effect on the fermentation characteristics and protein degradation of WGTG silage.

• Asian-Australasian Journal of Animal Sciences
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• v.12 no.8
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• pp.1205-1214
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• 1999

Whole lupinus albus seeds were pressure toasted at temperatures of 100, 118 and $136^{\circ}C$ for 3, 7, 15 and 30 min to study rumen degradation and post-rumen digestion and to determine optimal heating conditions for the Dutch dairy feed industry. In sacco nylon bag and mobile bag techniques were employed for rumen and intestine incubations to determine ruminal degradation characteristics and intestinal digestion of crude protein (CP) in 4 lactation rumen cannulated and 4 lactating intestinal cannulated Dutch dairy cows fed 47% hay and 53% concentrate according to Dutch dairy requirements. Measured rumen degradation characteristics were soluble fraction (S), undegradable fraction (U), potentially degradable fraction (D), lag time (T0) and rate of degradation (Kd) of insoluble but degradable fraction. Percentage bypass feed protein (BCP), ruminal microbial protein synthesized based on available nitrogen (N_MP) and that based on available energy (E_MP), true protein supplied to the small intestine (TPSI), truly absorbed BCP (ABCP), absorbed microbial protein (AVP) in the small intestine, endogenous protein losses in the digestion (ENDP), true digested protein in the small intestine (TAP or DVE in Dutch) and degraded protein balance (PDB or OEB in Dutch) were totally evaluated using the new Dutch DVE/OEB System. Pressure toasting decreased (p<0.001) rumen degradability of CP. It reduced S (p<0.05) and Kd (p=0.06), increased D (p<0.05) and U (p<0.01) but did not alter T0 (p>0.05), thus resulting in dramatically increased BCP (p<0.001) with increasing time and temperature from 73.7 (raw) up to 182.5 g/kg DM ($136^{\circ}C/15min$). Although rumen microbial protein synthesized based on available energy (E_MP) was reduced, true protein (microbial and bypass feed protein) supplied to the small intestine (TPSI) was increased (p<0.001) from 153.1 (raw) to 247.6 g/kg DM ($136^{\circ}C/15min$). Due to digestibility of BCP in the intestine not changing (p>0.05) average 87.8%, the absorbed BCP increased (p<0.001) from 62.3 (raw) to 153.7 g/kg DM ($136^{\circ}C/15min$). Therefore DVE value of true digested protein in the small intestine was significantly increased (p<0.001) from 118.9 (raw) to 197.0 g/kg DM ($136^{\circ}C/15min$) and OEB value of degraded protein balance was significantly reduced (p<0.001) from 147.2 (raw) to 63.1 g/kg DM ($136^{\circ}C/15min$). It was concluded that pressure toasting was effective in shifting degradation of CP of lupinus albus from the rumen to small intestine without changing intestinal digestion. Further studies are required on the degradation and digestion of individual amino acids and on the damaging effects of processing on amino acids, especially the first limiting amino acids.

• Asian-Australasian Journal of Animal Sciences
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• v.13 no.4
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• pp.535-542
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• 2000

Canola meal and seed are poor sources of ruminal undegraded protein (RUP). On average, canola meal and canola seed contains 35 and 14% RUP, respectively. Several protection methods are effective in reducing ruminal degradation of canola protein and in increasing RUP without affecting total tract protein digestibility. Heat (e.g., dry heat, moist heat and jet-sploding) and chemical (e.g., formaldehyde) treatments are the most common methods used to reduce ruminal degradability of canola protein. In most cases, heat treatments were found to be more effective than chemical treatments in protecting canola protein form ruminal degradation. Despite improvement in RUP content and intestinal availability of RUP, data form several studies showed little or no improvement in animal performance as a result of increasing the RUDP level of canola meal and seed.

• BMB Reports
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• v.51 no.10
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• pp.484-485
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• 2018

Receptor-interacting protein kinase-3 (RIP3 or RIPK3) is a serine-threonine kinase largely essential for necroptotic cell death; it also plays a role in some inflammatory diseases. High levels of RIP3 are likely sufficient to activate necroptotic and inflammatory pathways downstream of RIP3 in the absence of an upstream stimulus. For example, we have previously detected high levels or RIP3 in the skin of Toxic Epidermal Necrolysis patients; this correlates with increased phosphorylation of MLKL found in these patients. We have long surmised that there are molecular mechanisms to prevent anomalous activity of the RIP3 protein, and so prevent undesirable cell death and inflammatory effects when inappropriately activated. Recent discovery that Carboxyl terminus of Hsp 70-Interacting Protein (CHIP) could mediate ubiquitylation- and lysosome-dependent RIP3 degradation provides a potential protein that has this capacity. However, while screening for RIP3-binding proteins, we discovered that pellino E3 ubiquitin protein ligase 1 (PELI1) also interacts directly with RIP3 protein; further investigation in this study revealed that PELI1 also targets RIP3 for proteasome-dependent degradation. Interestingly, unlike CHIP, which targets RIP3 more generally, PELI1 preferentially targets kinase active RIP3 that has been phosphorylated on T182, subsequently leading to RIP3 degradation.

• Proceedings of the Korean Biophysical Society Conference
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• 2003.06a
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• pp.34-34
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• 2003

Protein unfolding is a key step in several cellular processes, including protein translocation across some membranes and protein degradation by ATP-dependent proteases. C1pAP protease and the proteasome can actively unfold proteins in a process that hydrolyzes ATP, These proteases catalyze unfolding by processively unraveling their substrates from the attachment point of the degradation signal. As a consequence, the ability of a protein to be degraded depends on its structure as well as its stability. An ${\alpha}$-helix is easier to unravel than a ${\beta}$-strand. In multidomain proteins, independently stable domains are unfolded sequentially. The steric constraints imposed on substrate proteins during their degradation by the proteasome were investigated by constructing a model protein in which specific parts of the polypeptide chain were covalently connected through disulfide bridges. The cross-linked model proteins were fully degraded by the proteasome, but two or more cross-links retarded the degradation slightly. Our results suggest that the pore of the proteasome allows the concurrent passage of at least three stretches of a polypeptide chain, and also explain the limited degradation by the proteasome that occurs in the processing of the transcription factor NF-KB, and also implicate difficulty in degradation of amyloidal aggregates by the proteasome

• Asian-Australasian Journal of Animal Sciences
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• v.13 no.10
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• pp.1377-1387
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• 2000

Two laboratory techniques: (1) an in vitro method with two procedures for measuring protein degradabilities and (2) an in vitro method with three procedures for measuring protein solubility, were investigated to determine which laboratory techniques could most accurately predict the quantity of rumen protein degradation kinetics of legume seeds after dry roasting under various conditions, in terms of (1) rumen protein disappearance ($D_j$, where j=0, 2, 4, 8, 12, 24 and 48 h incubation), (2) rumen protein effective degradability (EDCP), (3) the parameters describing rumen degradation characteristics (the soluble fraction: S, the potentially degradable fraction: D, undegradable fraction: U, lag time: T0 and the degradation rate: Kd) and (4) rumen bypass protein (BCP), which were determined by the method accepted internationally at present, in sacco nylon bag technique using the standardized Dutch method. Feeds evaluated were the raw and dry roasted whole faba (Vicia faba) beans (WFB) and whole lupin (Lupinus albus) seeds (WLS), each was dry roasted under various conditions (at 110, 130 or $150^{\circ}C$ for 15, 30 or 45 min). In vitro protein degradability ($D_1$_Auf and $D_{24}$_Auf) were determined using the modified Aufr re method by enzymatic hydrolysis for 1 h and 24 h using a protease extracted from Streptomyces griseus in a borate-phosphate buffer. In vitro protein solubility ($bf_1$_S, $bf_2$_S, $bf_3$_S) was measured in a borate-phosphate buffer with three different procedures. Results from laboratory techniques (in vitro) were correlated and linearly regressed with in sacco results. Of the three procedures of in vitro protein solubility evaluated, none of them could predict in sacco results with good precision. The highest Pearson correlation coefficient ($R^2$) was less than 0.50. Of two procedures of in vitro protein degradability studied, the $D_1$_Auf values were closely correlated with in sacco parameters: Kd, EDCP and %BCP with high R' values: 0.82, 0.85 and 0.85, respectively, and closely correlated with in sacco $D_j$ at 2, 4, 8 and 12 h rumen incubation with high $R^2$ values: 0.83, 0.91, 0.93 and 0.83, respectively. The $D_{24}$_Auf values could not predict in sacco results. The highest $R^2$ value was less then 0.40. These results indicated that in vitro protein solubility measured in borate-phosphate failed to identify differences in the rate and extent of protein degradation of legume seeds after dry roasting under various conditions and thus should not be used to predict rumen degradation, particularly for heat processed feedstuffs. But in vitro protein degradability using the modified Aufr re method by enzymatic hydrolysis for 1 h or possibly an intermediate time (>1 h and <24 h) is a promising laboratory procedure to detect effectiveness of dry roasting legume seeds on rumen protein degradation characteristics and could be used as a simple laboratory method to predict the rate and extent of protein degradation in the rumen in sacco with high accuracy. The equations to predict EDCP, Kd and BCP of dry roasted legume seeds (WLS and WFB) under various conditions are as follow: For both: EDCP (%)=-1.37+1.06*$D_1$_Auf ($R^2=0.85$, p<0.01). For both: Kd (%/h)=-21.81+0.49*$D_1$_Auf ($R^2=0.82$, p<0.01). For both: %BCP=103.37-1.07*$D_1$_Auf ($R^2=0.85$, p<0.01).

• The Korean Journal of Physiology and Pharmacology
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• v.24 no.6
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• pp.481-492
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• 2020

The present study aimed to examine the effect of allyl isothiocyanate (AITC) on chronic obstructive pulmonary disease and to investigate whether upregulation of multidrug resistance-associated protein 1 (MRP1) associated with the activation of the PARK7 (DJ-1)/nuclear factor erythroid 2-related factor 2 (Nrf2) axis. Lung function indexes and histopathological changes in mice were assessed by lung function detection and H&E staining. The expression levels of Nrf2, MRP1, heme oxygenase-1 (HO-1), and DJ-1 were determined by immunohistochemistry, Western blotting and reverse transcription-quantitative polymerase chain reaction. Next, the expression of DJ-1 in human bronchial epithelial (16HBE) cells was silenced by siRNA, and the effect of DJ-1 expression level on cigarette smoke extract (CSE)-stimulated protein degradation and AITC-induced protein expression was examined. The expression of DJ-1, Nrf2, HO-1, and MRP1 was significantly decreased in the wild type model group, while the expression of each protein was significantly increased after administration of AITC. Silencing the expression of DJ-1 in 16HBE cells accelerated CSE-induced protein degradation, and significantly attenuated the AITC-induced mRNA and protein expression of Nrf2 and MRP1. The present study describes a novel mechanism by which AITC induces MRP1 expression by protecting against CS/CSE-mediated DJ-1 protein degradation via activation of the DJ-1/Nrf2 axis.