| Field | Specification |
|---|---|
| Mfr No | |
| Activity | |
| Alternative Names | P1234;Non-structural polyprotein |
| Conjugate | |
| Endotoxin Level | |
| Expression System | |
| Form | Liquid or Lyophilized powder |
| Molecular Weight | |
| Product Type | |
| Protein Length | |
| Purity | |
| Reconstitution | |
| Species | |
| Storage | |
| Target | |
| UniProt # |
Overview
Recombinant Semliki forest virus Polyprotein P1234, partial is a recombinant protein preparation from Semliki forest virus (SFV) designed for use in assay development, binding studies, and functional characterization. Key attributes such as expression system, expressed region, and affinity tag(s) help researchers match the reagent to specific experimental readouts.
Key elements and design rationale
- Expression system: Baculovirus expression is commonly used for rapid, scalable production. For targets that require glycosylation or other post-translational modifications, consider how a prokaryotic system may affect folding or activity.
- Expression region: The expressed fragment (29-260aa) focuses the reagent on a defined domain/segment, which can influence binding interfaces and epitope availability.
- Tag(s)/format: His tags can support purification and detection in pull-down or binding assays; confirm that the tag position does not interfere with the interaction of interest.
- Purity: ≥90% (SDS-PAGE) provides a quick checkpoint for reagent quality in downstream analytical workflows.
- Form: Supplied as Liquid or Lyophilized powder; select the format that best fits your lab’s handling and aliquoting preferences.
Recombinant design choices (expression host, fragment boundaries, and tag configuration) help balance yield, solubility, and assay compatibility. Choose conditions and controls that match the recombinant format to your experimental question.
Biological background
SFV-P1234 has been reported to be involved in Inactive precursor of the viral replicase, which is activated by cleavages carried out by the viral protease nsP2. ; [Polyprotein P123]: The early replication complex formed by the polyprotein P123 and nsP4 synthesizes minus-strand RNAs. As soon P123 is cleaved into mature proteins, the plus-strand RNAs synthesis begins. ; [Polyprotein P123']: The early replication complex formed by the polyprotein P123' and nsP4 synthesizes minus-strand RNAs (Probable). Polyprotein P123' is a short-lived polyprotein that accumulates during early stage of infection (Probable). As soon P123' is cleaved into mature proteins, the plus-strand RNAs synthesis begins (Probable). ; [mRNA-capping enzyme nsP1]: Cytoplasmic capping enzyme that catalyzes two virus-specific reactions: methyltransferase and nsP1 guanylyltransferase. mRNA-capping is necessary since all viral RNAs are synthesized in the cytoplasm, and host capping enzymes are restricted to the nucleus (Probable). The enzymatic reaction involves a covalent link between 7-methyl-GMP and nsP1, whereas eukaryotic capping enzymes form a covalent complex only with GMP (Probable). nsP1 capping consists in the following reactions: GTP is first methylated into 7-methyl-GMP and then is covalently linked to nsP1 to form the m7GMp-nsP1 complex from which 7-methyl-GMP complex is transferred to the mRNA to create the cap structure (Probable). NsP1 is also needed for the initiation of the minus-strand RNAs synthesis. Probably serves as a membrane anchor for the replication complex composed of nsP1-nsP4 (Probable). Palmitoylated nsP1 is remodeling host cell cytoskeleton, and induces filopodium-like structure formation at the surface of the host cell. ; [Protease nsP2]: Multifunctional protein whose N-terminus is part of the RNA polymerase complex and displays NTPase, RNA triphosphatase and helicase activities. NTPase and RNA triphosphatase are involved in viral RNA capping and helicase keeps a check on the dsRNA replication intermediates (Probable). The C-terminus harbors a protease that specifically cleaves and releases the mature proteins. Required for the shutoff of minus-strand RNAs synthesis. Specifically inhibits the host IFN response by promoting the nuclear export of host STAT1. Also inhibits host transcription by inducing rapid proteasome-dependent degradation of POLR2A, a catalytic subunit of the RNAPII complex. The resulting inhibition of cellular protein synthesis serves to ensure maximal viral gene expression and to evade host immune response (Probable). ; [Non-structural protein 3']: Seems to be essential for minus-strand RNAs and subgenomic 26S mRNAs synthesis. Displays mono-ADP-ribosylhydrolase activity (Probable). ADP-ribosylation is a post-translational modification that controls various processes of the host cell and the virus probably needs to revert it for optimal viral replication (Probable). Binds proteins of FXR family and sequesters them into the viral RNA replication complexes thereby inhibiting the formation of host stress granules on viral mRNAs (Probable). The nsp3'-FXR complexes bind viral RNAs and probably orchestrate the assembly of viral replication complexes, thanks to the ability of FXR family members to self-assemble and bind DNA (Probable). ; [Non-structural protein 3]: Seems to be essential for minus-strand RNAs and subgenomic 26S mRNAs synthesis. Displays mono-ADP-ribosylhydrolase activity. ADP-ribosylation is a post-translational modification that controls various processes of the host cell and the virus probably needs to revert it for optimal viral replication. Binds proteins of G3BP family and sequesters them into the viral RNA replication complexes thereby inhibiting the formation of host stress granules on viral mRNAs. The nsp3-G3BP complexes bind viral RNAs and probably orchestrate the assembly of viral replication complexes, thanks to the ability of G3BP family members to self-assemble and bind DNA. ; [RNA-directed RNA polymerase nsP4]: RNA dependent RNA polymerase. Replicates genomic and antigenomic RNA by recognizing replications specific signals. The early replication complex formed by the polyprotein P123 and nsP4 synthesizes minus-strand RNAs. The late replication complex composed of fully processed nsP1-nsP4 is responsible for the production of genomic and subgenomic plus-strand RNAs.. When interpreting results, consider species context, domain architecture, and whether the recombinant format represents full-length or a defined region.
Research relevance and current trends
- Antigen and virulence-factor studies that compare strain- or domain-specific binding and immune recognition.
- Use of recombinant proteins as standards for quantitative assays and serology-oriented method development.
Common research applications
- Binding and interaction assays: quantify partner binding and rank conditions using plate-based formats or biophysical methods (SPR/BLI).
- Enzymology: assess catalytic activity and compare substrate preferences or inhibitor effects using appropriate controls.
- Assay development: use as a standard, spike-in control, or positive control where consistent specifications are required.
Interpretation typically relies on relative comparisons (treated vs control, mutant vs wild-type, or dose/time series) using consistent sample handling and appropriate normalization.
Notes for experimental interpretation
- Post-translational modifications: expression system can affect glycosylation and processing; interpret differences cautiously when comparing to native protein.
- Isoforms and domains: expressed regions may not capture all isoform-specific features; match fragment boundaries to your assay’s binding site.
- Controls: include blank matrix controls, tag-only controls (where relevant), and orthogonal readouts (e.g., WB/qPCR/ELISA) to support interpretation.
What is protein expression and purification?
Why is there no/low protein expression?
b. Rare codons. You should optimize codons, use strains supplementing rare codons, induce at lower temperature or grow in poor media.
c. Protein toxicity. You should use promoters with tighter regulation or lower plasmid copy number. Use pLysS/pLysE bearing strains in T7-based systems or strains that are better for the expression of toxic proteins. Start induction at high OD and shorten induction time. Add glucose when using expression vectors containing lac-based promoters.
How to avoid inclusion bodies and improve soluble expression?
b. Incorrect disulfide bond formation. You should add fusion partners, including thioredoxin, DsbA, DsbC. Clone in a vector containing secretion signal peptide to cell periplasm. Use gamiB (DE3)strains with oxidative cytoplasmic environment. Lower inducer concentration and induction temperature.
c. Incorrect folding. You should use a fusion partner. Co-express with molecular chaperones. Use strains with cold-adapted chaperones. Supplement media with chemical chaperones and cofactors. Reduce the inducer concentration and add fresh media. Induce for a shorter time at low temperature.
Why is the molecular weight of protein smaller than the predicted?
b. Imbalanced translation process of fusion protein. You should change another fusion tag or move fusion tag to C-terminal. You should induce for a shorter time at low temperature or change to poor media.
c. Protein degradation. You should replace specific protease sites. Use protease deficient strains. Induce at high OD. You should induce for a shorter time at low temperature or use protease inhibitors when breaking cells.
Why is the actual band size different from the predicted?
b. Post-translational cleavage. Many proteins are synthesized as pro-proteins, and then cleaved to give the active form.
c. Splice variants. Alternative splicing may create different sized proteins from the same gene.
d. Relative charge. The composition of amino acids have different relative charge which will affect the electrophoretic mobility.
e. Multimers such as dimerisation of a protein. This is usually prevented in reducing conditions, although strong interactions can result in the appearance of higher bands.
f. Protein structure such as disulfide bond, protein secondary structure or protein 3D structure formation.
g. Hydrophobic proteins, such as transmembrane proteins, may have difficulties in migrating into the gel, and thus resulting in different multi-banded patterns.
How to express a protein with bioactivity? Why is the protein inactive?
a. Low solubility of the protein. You should fuse desired protein to a fusion partners and lower temperature.
b. Lack of essential post translational modification. You should change another expression system.
c. Incomplete folding. You should use a fusion partner and use strains with cold-adapted chaperones. Co-express with molecular chaperones at lower temperature. Monitor disulfide bond formation and allow further folding in vitro.
d. Mutations in cDNA. You should sequence plasmid before and after induction or use a recA− strain to ensure plasmid stability. Transform E. coli before each expression round.
Why are our protein products almost invisible in pipes?
Tips: Before opening the lid, we recommend to centrifuge in a small centrifuge for 20-30 seconds firstly to ensure that the contents are on the bottom of the tube. Our quality control steps ensure that the amount of protein contained in each tube is accurate, although sometimes you can’t see the protein powder, but the protein content in the tube is still very accurate.
How is the protein purified? Is the purity guaranteed?
Although we guarantee a minimum purity standard of >85%, some of the proteins we prepared have a purity of 95% or even 97%.
How should I reconstitute and store the products?
As for short-term storage or usage, please use sterile deionized water to completely reconstitute proteins to 0.1-1.0 mg/mL. Aliquot after 10-15 minutes if needed and store at 4℃.
As for long-term storage, the cytokines or recombinant proteins are recommended to add 5-50% of glycerol (final concentration) and aliquot for long-term storage at -20℃/-80℃. Our default final concentration of glycerol is 50%. Customers could use it as reference.
What types of tags do you use for fusion?
What is the impact of a given tag type and any potential biological activity of the protein?
Can you remove the endotoxin?
Can you offer aseptic manufacture processing?
How to determine species cross-reactivity of cytokines?
b. Many mouse cytokines may also have effect on human cells, however, the activity may be lower than the corresponding human cytokines.
c. One of the few human cytokines will be more active than corresponding mouse cytokines when acting on mouse cells, such as IL-7.
d. Interferon, GM-CSF, IL-3 and IL-4 and other cytokines are species-specific and almost have no activity on non-homologous cells.
e. In contrast, fibroblast growth factor (FGF) and neurotrophin are highly conserved and both have good activity on cells of different species.
What is the general preservative? Which kind of preservative do you usually add?
What is the general protectant? What kind of protectant do you usually add?
Can’t Find What You’re Looking For? We can help you source the best match or customize a recombinant protein solution for your study. Options may include species (human/mouse/rat), protein region/domain (full-length vs fragment), tag or label (His/GST/FLAG/biotin/fluorescent), expression system (E. coli/HEK293/insect), purity grade, formulation (buffer, carrier-free, glycerol-free), activity/functional validation (binding or enzymatic assays), endotoxin level (low-endotoxin for cell-based work), mutants/variants (point mutations, isoforms), and bulk or custom packaging. Click Talk to a Scientist to submit a request form, email us at support@biohippo.com, or explore our Research Services for additional support. Our team will be in contact with you shortly.
