Homoserine dehydrogenase (HSD), encoded by the hom gene, is a key enzyme within the aspartate pathway, which reversibly catalyzes the conversion of l-aspartate β-semialdehyde to l-homoserine (l-Hse), utilizing both NAD(H) or NADP(H) as a coenzyme.
On this work, we introduced the primary characterization of the HSD from the symbiotic Polynucleobacter necessaries subsp. necessarius (PnHSD) produced in Escherichia coli. Sequence evaluation confirmed that PnHSD is an ACT domain-containing monofunctional HSD with 436 amnio acid residues.
SDS-PAGE and Western blot demonstrated that PnHSD could possibly be overexpressed in E. coli BL21(DE3) cell as a soluble type through the use of SUMO fusion approach. It could possibly be purified to obvious homogeneity for biochemical characterization.
Dimension-exclusion chromatography revealed that the purified PnHSD has a local molecular mass of ∼160 kDa, indicating a homotetrameric construction. The oxidation exercise of PnHSD was studied on this work. Kinetic evaluation revealed that PnHSD displayed an as much as 1460-fold choice for NAD+ over NADP+, in distinction to its homologs.
The purified PnHSD displayed maximal exercise at 35 °C and pH 11. Much like its NAD+-dependent homolog, neither NaCl and KCl activation nor L-Thr inhibition on the enzymatic exercise of PnHSD was noticed. These outcomes will contribute to a greater understanding of the coenzyme specificity of the HSD household and the aspartate pathway of P. necessarius.
Secondary Construction of Human De Novo Advanced Gene Product NCYM Analyzed by Vacuum-Ultraviolet Round Dichroism
NCYM, a cis-antisense gene of MYCN, encodes a Homininae-specific protein that promotes the aggressiveness of human tumors. Newly advanced genes from non-genic areas are referred to as de novo genes, and NCYM was the primary de novo gene whose oncogenic features had been validated in vivo.
Concentrating on NCYM utilizing medicine is a possible technique for most cancers remedy; nevertheless, the NCYM construction have to be decided earlier than drug design. On this examine, we employed vacuum-ultraviolet round dichroism to guage the secondary construction of NCYM.
The SUMO–tagged NCYM and the remoted SUMO tag in each hydrogenated and perdeuterated varieties had been synthesized and purified in a cell-free in vitro system, and vacuum-ultraviolet round dichroism spectra had been measured. Vital variations between the tagged NCYM and the remoted tag had been evident within the wavelength vary of 190-240 nm.
The round dichroism spectral information mixed with a neural community system enabled to foretell the secondary construction of NCYM on the amino acid stage. The 129-residue tag consists of α-helices (roughly 14%) and β-strands (roughly 29%), which corresponded to the values calculated from the atomic construction of the tag.
The 238-residue tagged NCYM contained roughly 17% α-helices and 27% β-strands. The situation of the secondary construction predicted utilizing the neural community revealed that these secondary constructions had been enriched within the Homininae-specific area of NCYM.
Deuteration of NCYM altered the secondary construction at D90 from an α-helix to a different construction aside from α-helix and β-strand though this alteration was inside the experimental error vary. All 4 nonsynonymous single-nucleotide polymorphisms (SNPs) in human populations had been on this area, and the amino acid alteration in SNP N52S enhanced Myc-nick manufacturing.
The D90N mutation in NCYM promoted NCYM-mediated MYCN stabilization. Our outcomes reveal the secondary construction of NCYM and demonstrated that the Homininae-specific area of NCYM is liable for MYCN stabilization.
Enhancement of Solubility, Purification, and Inclusion Physique Refolding of Energetic Human Mitochondrial Aldehyde Dehydrogenase 2
Mitochondrial aldehyde dehydrogenase 2 (ALDH2) is predominantly linked with acetaldehyde detoxing within the second stage of alcohol metabolism. To intensively examine ALDH2 perform, a better purity and uniform composition of the protein is required.
An environment friendly Escherichia coli system for ALDH2 expression was developed through the use of His and a small ubiquitin-related modifier fusion tag. Many of the recombinant ALDH2s had been expressed within the type of inclusion our bodies.
The ALDH2-enriched inclusion our bodies had been denatured with 6 M guanidine hydrochloride, after which ALDH2 was ultrafitrated. Lastly, ALDH2 was efficiently purified by affinity and gel filtration chromatography. The purified ALDH2 was lastly preserved by the vacuum freeze-drying technique, and its purity was decided to be larger than 95%, with a closing media yield of 33.89 mg/L.
The particular exercise of ALDH2 was 6.1 × 104 U/mg. This work was the primary to report pET-SUMO-ALDH2 recombinant plasmid expression in Escherichia coli, and the inclusion our bodies had been remoted and refolded. Lastly, the purified ALDH2 had comparatively larger purity, yield, and organic exercise.
Toxin Fused with SUMO Tag: A New Expression Vector Technique to Acquire Recombinant Venom Toxins with Simple Tag Removing contained in the Micro organism.
Many animal toxins could goal the identical molecules that must be managed in sure pathologies; subsequently, some toxins have led to the formulation of medicine which can be presently used, and lots of different medicine are nonetheless beneath improvement. However, amassing ample toxins from the unique supply could be a limiting think about finding out their organic actions.
Thus, molecular biology methods have been utilized with the intention to get hold of giant quantities of recombinant toxins into Escherichia coli. Nevertheless, most animal toxins are tough to precise on this system, which leads to insoluble, misfolded, or unstable proteins.
To unravel these points, toxins have been fused with tags that will enhance protein expression, solubility, and stability. Amongst these tags, the SUMO (small ubiquitin-related modifier) has been proven to be very environment friendly and could be eliminated by the Ulp1 protease.
Nevertheless, eradicating SUMO is a labor- and time-consuming course of. To boost this method, right here we present the development of a bicistronic vector that permits the expression of any protein fused to each the SUMO and Ulp1 protease.
On this approach, after expression, Ulp1 is ready to cleave SUMO and depart the protein interest-free and prepared for purification. This technique was validated by the expression of a brand new phospholipase D from the spider Loxosceles gaucho and a disintegrin from the Bothrops insularis snake.
Each recombinant toxins confirmed good yield and preserved organic actions, indicating that the bicistronic vector could also be a viable technique to provide proteins which can be tough to precise.
A Mutant Sumo Facilitates Fast Plasmid Building for Expressing Proteins with Native N-termini After Tag Removing.
Sumo is without doubt one of the fusion tags generally used to reinforce the expression and the solubility of recombinant proteins. One benefit of utilizing sumo is that the elimination of the sumo tag is very particular as a result of its recognition by a sumo protease is decided by its structural traits, as an alternative of the sequence of a brief peptide.
Just lately, it was reported that sumo may be used as a protease recognition web site to facilitate the elimination of different fusion tags, equivalent to MBP, when sumo itself shouldn’t be appropriate to reinforce the solubility of a specific goal protein.
Utilizing sumo as a recognition web site is very fascinating when the goal protein must have its native N terminus. Nevertheless, establishing such a plasmid entails a couple of cloning step as a result of the N terminus of the goal protein must be the subsequent residue after the diglycine of sumo.
Recombinant Human NEDD8 (N-6His, SUMO tag) |
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| CG48-50ug | Novoprotein | 50ug | EUR 114 |
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Description: Lyophilized from a 0.2 μm filtered solution of 20mM PB，150mM NaCl，5% Trehalose,pH7.4. |
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Recombinant Human Ubiquitin-Conjugating Enzyme E2 K/UBE2K (N-6His, SUMO tag) |
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| CG45-10ug | Novoprotein | 10ug | EUR 96 |
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Description: Supplied as a 0.2 μm filtered solution of 20mM PB,150mM NaCl,pH7.4. |
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Recombinant Human Ubiquitin-Conjugating Enzyme E2 K/UBE2K (N-6His, SUMO tag) |
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| CG45-1mg | Novoprotein | 1mg | EUR 1216.8 |
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Description: Supplied as a 0.2 μm filtered solution of 20mM PB,150mM NaCl,pH7.4. |
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Recombinant Human Ubiquitin-Conjugating Enzyme E2 K/UBE2K (N-6His, SUMO tag) |
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| CG45-500ug | Novoprotein | 500ug | EUR 790.8 |
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Description: Supplied as a 0.2 μm filtered solution of 20mM PB,150mM NaCl,pH7.4. |
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Recombinant Human Ubiquitin-Conjugating Enzyme E2 K/UBE2K (N-6His, SUMO tag) |
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| CG45-50ug | Novoprotein | 50ug | EUR 169.2 |
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Description: Supplied as a 0.2 μm filtered solution of 20mM PB,150mM NaCl,pH7.4. |
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SUMO3 protein (His tag) |
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| 80R-1512 | Fitzgerald | 100 ug | EUR 366 |
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Description: Purified recombinant Human SUMO3 protein |
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SUMO1 protein (His tag) |
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| 80R-2035 | Fitzgerald | 100 ug | EUR 386.4 |
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Description: Recombinant human SUMO1 protein (His tag) |
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pCold- SUMO |
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| PVT0606 | Lifescience Market | 2 ug | EUR 390 |
pET28a- SUMO |
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| PVT0080 | Lifescience Market | 2 ug | EUR 390 |
Sumo Peptide |
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| 3969P | ProSci | 0.05 mg | EUR 197.7 |
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Description: (NT) Sumo peptide |
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Sumo Antibody |
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| 24467-100ul | SAB | 100ul | EUR 468 |
Sumo Antibody |
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| 3969-002mg | ProSci | 0.02 mg | EUR 206.18 |
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Description: Sumo Antibody: The sumo family of proteins is related both structurally and functionally to ubiquitin in that they are post-translationally attached to the e-amino group of a lysine residue of the substrate protein. This sumoylation plays a number of roles in DNA replication and repair, protein targeting to various subnuclear structures, and the regulation of numerous cellular processes including the inflammatory response in mammalian cells. Sumo was initially identified as a covalent modification of RanGAP1 in studies on nuclear import in mammalian cells. More recently, sumo has been shown to be involved in the regulation of transcription factors, possibly by enhancing their interactions with co-repressors. Sumo is also thought to play some role in the modulation of ubiquitin-mediated degradation of proteins by acting as an inhibitor. At least four different isoforms of sumo are known to exist; Sumo antibody will only recognize isoform 1. |
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Sumo Antibody |
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| 3969-01mg | ProSci | 0.1 mg | EUR 523.7 |
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Description: Sumo Antibody: The sumo family of proteins is related both structurally and functionally to ubiquitin in that they are post-translationally attached to the e-amino group of a lysine residue of the substrate protein. This sumoylation plays a number of roles in DNA replication and repair, protein targeting to various subnuclear structures, and the regulation of numerous cellular processes including the inflammatory response in mammalian cells. Sumo was initially identified as a covalent modification of RanGAP1 in studies on nuclear import in mammalian cells. More recently, sumo has been shown to be involved in the regulation of transcription factors, possibly by enhancing their interactions with co-repressors. Sumo is also thought to play some role in the modulation of ubiquitin-mediated degradation of proteins by acting as an inhibitor. At least four different isoforms of sumo are known to exist; Sumo antibody will only recognize isoform 1. |
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SUMO Antibody |
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| F53483-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 322.15 |
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Description: Covalent modification of target lysines by SUMO (small ubiquitin-like modifier) modulates processes such as protein localization, transcription, nuclear transport, mitosis, DNA replication and repair, signal transduction, and viral reproduction. SUMO does not seem to be involved in protein degradation and may in fact function as an antagonist of ubiquitin in the degradation process. In the development of Drosophila, SUMO plays a maternal role in anterior-posterior (A/P) polarity and patterning. |
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SUMO Antibody |
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| F40023-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 322.15 |
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Description: Covalent modification of target lysines by SUMO (small ubiquitin-like modifier) modulates processes such as protein localization, transcription, nuclear transport, mitosis, DNA replication and repair, signal transduction, and viral reproduction. SUMO does not seem to be involved in protein degradation and may in fact function as an antagonist of ubiquitin in the degradation process. In the development of Drosophila, SUMO plays a maternal role in anterior-posterior (A/P) polarity and patterning. |
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SUMO Antibody |
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| F42008-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 322.15 |
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Description: Ubiquitin-like protein that can be covalently attached to proteins as a monomer or a lysine-linked polymer. Covalent attachment via an isopeptide bond to its substrates requires prior activation by the E1 complex SAE1-SAE2 and linkage to the E2 enzyme UBE2I, and can be promoted by E3 ligases such as PIAS1-4, RANBP2 or CBX4. This post-translational modification on lysine residues of proteins plays a crucial role in a number of cellular processes such as nuclear transport, DNA replication and repair, mitosis and signal transduction. Involved for instance in targeting RANGAP1 to the nuclear pore complex protein RANBP2. Covalently attached to the voltage-gated potassium channel KCNB1; this modulates the gating characteristics of KCNB1. Polymeric SUMO1 chains are also susceptible to polyubiquitination which functions as a signal for proteasomal degradation of modified proteins. May also regulate a network of genes involved in palate development. [UniProt] |
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SUMO Antibody |
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| F42009-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 322.15 |
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Description: Covalent modification of target lysines by SUMO (small ubiquitin-like modifier) modulates processes such as protein localization, transcription, nuclear transport, mitosis, DNA replication and repair, signal transduction, and viral reproduction. |
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SUMO-1 Protein |
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| E28000 | EpiGentek | 100 µg | EUR 423.66 |
SUMO-2 Protein |
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| E28002 | EpiGentek | 100 µg | EUR 423.66 |
SUMO-1 Antibody |
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| 20-abx123448 | Abbexa |
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SUMO-1 Antibody |
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| 48765-100ul | SAB | 100ul | EUR 399.6 |
SUMO-1 Antibody |
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| 48765-50ul | SAB | 50ul | EUR 286.8 |
SUMO-2 Antibody |
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| F52344-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 330.65 |
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Description: Ubiquitin-like protein that can be covalently attached to proteins as a monomer or as a lysine-linked polymer. Covalent attachment via an isopeptide bond to its substrates requires prior activation by the E1 complex SAE1-SAE2 and linkage to the E2 enzyme UBE2I, and can be promoted by an E3 ligase such as PIAS1-4, RANBP2 or CBX4. This post-translational modification on lysine residues of proteins plays a crucial role in a number of cellular processes such as nuclear transport, DNA replication and repair, mitosis and signal transduction. Polymeric SUMO2 chains are also susceptible to polyubiquitination which functions as a signal for proteasomal degradation of modified proteins. |
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SUMO-2 Antibody |
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| F42016-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 322.15 |
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Description: SUMO2 and SUMO3 are members of the SUMO (small ubiquitin-like modifier) protein family. This protein family functions in a manner similar to ubiquitin in that it is bound to target proteins as part of a post-translational modification system. However, unlike ubiquitin which targets proteins for degradation, this protein is involved in a variety of cellular processes, such as nuclear transport, transcriptional regulation, apoptosis, and protein stability. In vertebrates, three members of the SUMO family have been described, SUMO 1 and the functionally distinct homologues SUMO 2 and SUMO 3. SUMO modification sites present in the N terminal regions of SUMO 2 and SUMO 3 are utilized by SAE1/SAE2 (SUMO E1) and Ubc9 (SUMO E2) to form polymeric chains of SUMO 2 and SUMO 3 on protein substrates, a property not shared by SUMO 1. |
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SUMO-3 Antibody |
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| F42017-0.08ML | NSJ Bioreagents | 0.08 ml | EUR 140.25 |
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Description: SUMO2 and SUMO3 are members of the SUMO (small ubiquitin-like modifier) protein family. This protein family functions in a manner similar to ubiquitin in that it is bound to target proteins as part of a post-translational modification system. However, unlike ubiquitin which targets proteins for degradation, this protein is involved in a variety of cellular processes, such as nuclear transport, transcriptional regulation, apoptosis, and protein stability. In vertebrates, three members of the SUMO family have been described, SUMO 1 and the functionally distinct homologues SUMO 2 and SUMO 3. SUMO modification sites present in the N terminal regions of SUMO 2 and SUMO 3 are utilized by SAE1/SAE2 (SUMO E1) and Ubc9 (SUMO E2) to form polymeric chains of SUMO 2 and SUMO 3 on protein substrates, a property not shared by SUMO 1. |
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SUMO-3 Antibody |
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| F42017-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 322.15 |
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Description: SUMO2 and SUMO3 are members of the SUMO (small ubiquitin-like modifier) protein family. This protein family functions in a manner similar to ubiquitin in that it is bound to target proteins as part of a post-translational modification system. However, unlike ubiquitin which targets proteins for degradation, this protein is involved in a variety of cellular processes, such as nuclear transport, transcriptional regulation, apoptosis, and protein stability. In vertebrates, three members of the SUMO family have been described, SUMO 1 and the functionally distinct homologues SUMO 2 and SUMO 3. SUMO modification sites present in the N terminal regions of SUMO 2 and SUMO 3 are utilized by SAE1/SAE2 (SUMO E1) and Ubc9 (SUMO E2) to form polymeric chains of SUMO 2 and SUMO 3 on protein substrates, a property not shared by SUMO 1. |
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SUMO-1 Antibody |
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| F43187-0.4ML | NSJ Bioreagents | 0.4 ml | EUR 322.15 |
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Description: This gene encodes a protein that is a member of the SUMO (small ubiquitin-like modifier) protein family. It functions in a manner similar to ubiquitin in that it is bound to target proteins as part of a post-translational modification system. However, unlike ubiquitin which targets proteins for degradation, this protein is involved in a variety of cellular processes, such as nuclear transport, transcriptional regulation, apoptosis, and protein stability. It is not active until the last four amino acids of the carboxy-terminus have been cleaved off. Several pseudogenes have been reported for this gene. Alternate transcriptional splice variants encoding different isoforms have been characterized. |
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SUMO-1 Antibody |
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| V9095-100UG | NSJ Bioreagents | 100 ug | EUR 349.3 |
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Description: This mAb is specific to SUMO-1 and shows no cross-reaction with either SUMO2 or SUMO3. The small ubiquitin-related modifier (SUMO) proteins, which include SUMO1, SUMO2 and SUMO3, belong to the ubiquitin-like protein family. Like ubiquitin, the SUMO proteins are synthesized is precursor proteins that undergo processing before conjugation to target proteins. Also, both utilize the E1, E2, and E3 cascade enzymes for conjugation. However, SUMO and ubiquitin differ with respect to targeting. Ubiquitination predominantly targets proteins for degradation, whereas sumoylation targets proteins to a variety of cellular processing, including nuclear transport, transcriptional regulation, apoptosis and protein stability. The unconjugated SUMO1 protein localizes to the nuclear membrane. |
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SUMO-1 Antibody |
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| V9095-20UG | NSJ Bioreagents | 20 ug | EUR 153.3 |
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Description: This mAb is specific to SUMO-1 and shows no cross-reaction with either SUMO2 or SUMO3. The small ubiquitin-related modifier (SUMO) proteins, which include SUMO1, SUMO2 and SUMO3, belong to the ubiquitin-like protein family. Like ubiquitin, the SUMO proteins are synthesized is precursor proteins that undergo processing before conjugation to target proteins. Also, both utilize the E1, E2, and E3 cascade enzymes for conjugation. However, SUMO and ubiquitin differ with respect to targeting. Ubiquitination predominantly targets proteins for degradation, whereas sumoylation targets proteins to a variety of cellular processing, including nuclear transport, transcriptional regulation, apoptosis and protein stability. The unconjugated SUMO1 protein localizes to the nuclear membrane. |
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SUMO-1 Antibody |
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| V9095IHC-7ML | NSJ Bioreagents | 7 ml | EUR 349.3 |
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Description: This mAb is specific to SUMO-1 and shows no cross-reaction with either SUMO2 or SUMO3. The small ubiquitin-related modifier (SUMO) proteins, which include SUMO1, SUMO2 and SUMO3, belong to the ubiquitin-like protein family. Like ubiquitin, the SUMO proteins are synthesized is precursor proteins that undergo processing before conjugation to target proteins. Also, both utilize the E1, E2, and E3 cascade enzymes for conjugation. However, SUMO and ubiquitin differ with respect to targeting. Ubiquitination predominantly targets proteins for degradation, whereas sumoylation targets proteins to a variety of cellular processing, including nuclear transport, transcriptional regulation, apoptosis and protein stability. The unconjugated SUMO1 protein localizes to the nuclear membrane. |
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SUMO-1 Antibody |
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| V9095SAF-100UG | NSJ Bioreagents | 100 ug | EUR 349.3 |
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Description: This mAb is specific to SUMO-1 and shows no cross-reaction with either SUMO2 or SUMO3. The small ubiquitin-related modifier (SUMO) proteins, which include SUMO1, SUMO2 and SUMO3, belong to the ubiquitin-like protein family. Like ubiquitin, the SUMO proteins are synthesized is precursor proteins that undergo processing before conjugation to target proteins. Also, both utilize the E1, E2, and E3 cascade enzymes for conjugation. However, SUMO and ubiquitin differ with respect to targeting. Ubiquitination predominantly targets proteins for degradation, whereas sumoylation targets proteins to a variety of cellular processing, including nuclear transport, transcriptional regulation, apoptosis and protein stability. The unconjugated SUMO1 protein localizes to the nuclear membrane. |
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SUMO-1(SUMO1/1188) Antibody |
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| BNC941188-100 | Biotium | 100uL | EUR 238.8 |
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Description: Primary antibody against SUMO-1(SUMO1/1188), CF594 conjugate, Concentration: 0.1mg/mL |
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SUMO-1(SUMO1/1188) Antibody |
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| BNC941188-500 | Biotium | 500uL | EUR 652.8 |
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Description: Primary antibody against SUMO-1(SUMO1/1188), CF594 conjugate, Concentration: 0.1mg/mL |
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SUMO-2(SUMO2/1199) Antibody |
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| BNC941199-100 | Biotium | 100uL | EUR 238.8 |
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Description: Primary antibody against SUMO-2(SUMO2/1199), CF594 conjugate, Concentration: 0.1mg/mL |
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SUMO-2(SUMO2/1199) Antibody |
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| BNC941199-500 | Biotium | 500uL | EUR 652.8 |
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Description: Primary antibody against SUMO-2(SUMO2/1199), CF594 conjugate, Concentration: 0.1mg/mL |
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Right here, we report the development of a brand new vector with a mutant sumo tag. The incorporation of a Pvu II web site close to the three’ finish of tag coding sequence permits fast building of plasmids for producing proteins with native termini. Its utilization consists of producing recombinant meals allergens for finding out conformational IgE epitopes.
