5,318 research outputs found
Control of protein quality and stoichiometries by N-terminal acetylation and the N-end rule pathway
No full textN-alpha-terminal acetylation of cellular proteins was recently discovered to create specific degradation signals termed Ac/N-degrons and targeted by the Ac/N-end rule pathway. We show that Hcn1, a subunit of the APC/C ubiquitin ligase, contains an Ac/N-degron that is repressed by Cut9, another APC/C subunit and the ligand of Hcn1. Cog1, a subunit of the Golgi-associated COG complex, is also shown to contain an Ac/N-degron. Cog2 and Cog3, direct ligands of Cog1, can repress this degron. The subunit decoy technique was used to show that the long-lived endogenous Cog1 is destabilized and destroyed via its activated (unshielded) Ac/N-degron if the total level of Cog1 increased in a cell. Hurl and Cog1 are the first examples of protein regulation through the physiologically relevant transitions that shield and unshield natural Ac/N-degrons. This mechanistically straightforward circuit can employ the demonstrated conditionality of Ac/N-degrons to regulate subunit stoichiometries and other aspects of protein quality control.X1111389sciescopu
Regulation of peptide import through phosphorylation of Ubr1, the ubiquitin ligase of the N-end rule pathway
Full text linkSubstrates of the N-end rule pathway include proteins with destabilizing N-terminal residues. These residues are recognized by E3 ubiquitin ligases called N-recognins. Ubr1 is the N-recognin of the yeast Saccharomyces cerevisiae. Extracellular amino acids or short peptides up-regulate the peptide transporter gene PTR2, thereby increasing the capacity of a cell to import peptides. Cup9 is a transcriptional repressor that down-regulates PTR2. The induction of PTR2 by peptides or amino acids involves accelerated degradation of Cup9 by the N-end rule pathway. We report here that the Ubr1 N-recognin, which conditionally targets Cup9 for degradation, is phosphorylated in vivo at multiple sites, including Ser(300) and Tyr(277). We also show that the type-I casein kinases Yck1 and Yck2 phosphorylate Ubr1 on Ser(300), and thereby make possible ("prime'') the subsequent (presumably sequential) phosphorylations of Ubr1 on Ser(296), Ser(292), Thr(288), and Tyr(277) by Mck1, a kinase of the glycogen synthase kinase 3 (Gsk3) family. Phosphorylation of Ubr1 on Tyr(277) by Mck1 is a previously undescribed example of a cascade-based tyrosine phosphorylation by a Gsk3-type kinase outside of autophosphorylation. We show that the Yck1/Yck2-mediated phosphorylation of Ubr1 on Ser(300) plays a major role in the control of peptide import by the N-end rule pathway. In contrast to phosphorylation on Ser(300), the subsequent (primed) phosphorylations, including the one on Tyr(277), have at most minor effects on the known properties of Ubr1, including regulation of peptide import. Thus, a biological role of the rest of Ubr1 phosphorylation cascade remains to be identified.open112324sciescopu
N-terminal acetylation of cellular proteins creates specific degradation signals
No full textThe retained N-terminal methionine (Met) residue of a nascent protein is often N-terminally acetylated (Nt-acetylated). Removal of N-terminal Met by Met-aminopeptidases frequently leads to Nt-acetylation of the resulting N-terminal alanine (Ala), valine (Val), serine (Ser), threonine (Thr), and cysteine (Cys) residues. Although a majority of eukaryotic proteins (for example, more than 80% of human proteins) are cotranslationally Nt-acetylated, the function of this extensively studied modification is largely unknown. Using the yeast Saccharomyces cerevisiae, we found that the Nt-acetylated Met residue could act as a degradation signal (degron), targeted by the Doa10 ubiquitin ligase. Moreover, Doa10 also recognized the Nt-acetylated Ala, Val, Ser, Thr, and Cys residues. Several examined proteins of diverse functions contained these N-terminal degrons, termed N-Ac-degrons, which are a prevalent class of degradation signals in cellular proteins.X11308291sciescopu
RECONSTRUCTION ON SI(100) SURFACES
No full textWe have observed several reconstructions on a Si(100) surface with different annealing procedures. The observed reconstructed phases are the coexistence of the (2X2) phase and the (2X8) phase after high-temperature (greater than or similar to 950 K) annealing followed by quenching, and the half-order streak with the presence of the (2X1) phase after low-temperature (less than or similar to 950 K) annealing. The phase transition from the metastable (2X2) and (2X8) phases to the stable half-order streak is reversible upon annealing temperature and cooling rate. The distribution of kinks and missing dimer defects is expected to be the main cause of these reconstructions.This work was supported by the Ability enhancement project at materials evaluation center, Koera Research Institute of standars and Science. Also, part of this work is supported by the center for molecular science at KAIST
Size effect on the fiber strength of composite pressure vessels
No full textIn this study, experimental tests and an analytical approach are conducted to verify the size effect on the fiber strength of a composite pressure vessel. As an analytical method, the Weibull weakest link model and the sequential multi-step failure model are considered and mutually compared. In the case of carbon fiber tensile strength, there is no large difference between the analytical methods for the volumetric size effect. To verify the validity of the analytical approach, experimental tests were performed using fiber strand specimens, unidirectional laminate specimens and composite pressure vessels. Good agreement for fiber strength distribution was shown between the test data and predicted results. The volumetric size effect shows the clearly observed tendency towards fiber strength degradation with increasing stressed volume. Because the volumetric size effect depends on material and processing factors, the reduction of fiber strength due to the stressed volume shows different values according to the variation of material and processing conditions
The N-end rule pathway is mediated by a complex of the RING-type Ubr1 and HECT-type Ufd4 ubiquitin ligases
No full textSubstrates of the N-end rule pathway are recognized by the Ubr1 E3 ubiquitin ligase through their destabilizing amino-terminal residues. Our previous work showed that the Ubr1 E3 and the Ufd4 E3 together target an internal degradation signal (degron) of the Mgt1 DNA repair protein. Ufd4 is an E3 enzyme of the ubiquitin-fusion degradation (UFD) pathway that recognizes an N-terminal ubiquitin moiety. Here we show that the RING-type Ubr1 E3 and the HECT-type Ufd4 E3 interact, both physically and functionally. Although Ubr1 can recognize and polyubiquitylate an N-end rule substrate in the absence of Ufd4, the Ubr1-Ufd4 complex is more processive in that it produces a longer substrate-linked polyubiquitin chain. Conversely, Ubr1 can function as a polyubiquitylation-enhancing component of the Ubr1-Ufd4 complex in its targeting of UFD substrates. We also found that Ubr1 can recognize the N-terminal ubiquitin moiety. These and related advances unify two proteolytic systems that have been studied separately for two decades.X117261sciescopu
Temperature forming of Ti-6Al-4V alloy considering microstructural evolution
No full textA study has been made to investigate the high temperature deformation behavior of Ti-6Al-4V alloy and to predict the final microstructure under given forming conditions. Equiaxed and Widmanstatten microstructures of Ti-6Al-4V alloys were prepared as initial microstructures. By performing the compression tests at high temperatures (700 similar to 1100degreesC) and a wide range of strain rates (10(4) similar to 10(2)/s), various parameters such as strain rate sensitivity (m).and activation energy (Q) were calculated and used to establish constitutive equations. When the specimens were deformed up to epsilon(c) approximate to 0.6, equiaxed microstructure did not show any significant changes in microstructure, while Widmanstatten microstructure revealed considerable flow softening, which was attributed to the globularization of a platelet at the temperature range of 800similar to970degreesC and at the strain rate range of 10(-4)similar to10(-2)/s. To predict the final microstructure after forming, finite element analysis was performed considering the microstructural evolution during the deformation. The grain size and the volume fraction of second phase of deformed body were predicted and compared with the experimental results.113sciescopu
Two proteolytic pathways regulate DNA repair by cotargeting the Mgt1 alkylguanine transferase
Full text linkO-6-methylguanine (O(6)meG) and related modifications of guanine in double-stranded DNA are functionally severe lesions that can be produced by many alkylating agents, including N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), a potent carcinogen. O(6)meG is repaired through its demethylation by the O-6-alkylguanine-DNA alkyltransferase (AGT). This protein is called Mgmt (or MGMT) in mammals and Mgt1 in the yeast Saccharomyces cerevisiae. AGT proteins remove methyl and other alkyl groups from an alkylated O-6 in guanine by transferring the adduct to an active-site cysteine residue. The resulting S-alkyl-Cys of AGT is not restored back to Cys, so repair proteins of this kind can act only once. We report here that S. cerevisiae Mgt1 is cotargeted for degradation, through a degron near its N terminus, by 2 ubiquitin-mediated proteolytic systems, the Ubr1/Rad6-dependent N-end rule pathway and the Ufd4/Ubc4-dependent ubiquitin fusion degradation (UFD) pathway. The cotargeting of Mgt1 by these pathways is synergistic, in that it increases not only the yield of polyubiquitylated Mgt1, but also the processivity of polyubiquitylation. The N-end rule and UFD pathways comediate both the constitutive and MNNG-accelerated degradation of Mgt1. Yeast cells lacking the Ubr1 and Ufd4 ubiquitin ligases were hyperresistant to MNNG but hypersensitive to the toxicity of overexpressed Mgt1. We consider ramifications of this discovery for the control of DNA repair and mechanisms of substrate targeting by the ubiquitin system.open1148sciescopu
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