Recombinant Human R-spondin 1, Animal-Free Protein

Catalog # Availability Size / Price Qty
Qk006-0050
Qk006-0500
Recombinant Human R-spondin 1, Animal-Free Protein Bioactivity
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Recombinant Human R-spondin 1, Animal-Free Protein Summary

Product Specifications

Endotoxin Level
<0.10 EU per 1 μg of the protein by the LAL method.
Activity
No significant difference between EC50 of reference and test lots
Source
E. coli-derived human R-Spondin 1 protein
Accession #
Predicted Molecular Mass
13 kDa
SDS-PAGE
Monomeric R-spondin 1 protein only

Product Datasheets

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QK006

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.

What formulation is right for me?

In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.

Qk006

Formulation Lyophilized from acetonitrile/TFA
Reconstitution Resuspend in 10mM HCl at >100 µg/ml, prepare single use aliquots, add carrier protein if desired.
Shipping The product is shipped lyophilized at ambient temperture, on ice blocks or dry ice. Shipping at ambient temperture does not affect the bioactivity or stability of the protein. Upon reciept, store immediately at the conditions stated below.
Stability & Storage: Store lyophilized protein between -20 and -80 °C until the date of expiry. Avoid freeze-thaw cycles.

Scientific Data

Bioactivity View Larger

R-spondin 1 activity is determined using the Wnt-responsive firefly luciferase reporter assay as it enhances Wnt-beta catenin signaling in HEK239T cells. HEK293T cells transfected with reporter TOP-FLASH are treated in triplicate with increasing concentration of R-spondin 1 (diluted in DMEM with 0.5 % of FCS), in the presence of Wnt-conditioned media (1:8 dilution). Cells are grown overnight, and luciferase activity is measured and normalized.EC50 = 75.5 ng/ml (5.8 nM).

SDS-PAGE View Larger

RSPO1 migrates as a single band at 16 kDa in non-reducing (NR) and 13 kDa in reducing (R) conditions.Purified recombinant protein (7 µg) was resolved using 15% w/v SDS-PAGE in reduced (+ beta -mercaptothanol, R) and non-reduced conditions (NR) and stained with Coomassie Brilliant Blue R250.

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Background: R-Spondin 1

R-Spondin 1 (RSPO1, Roof plate-specific Spondin 1), also known as cysteine-rich and single thrombospondin domain containing protein 3 (Cristin 3), is a 27 kDa secreted protein that shares ~40% amino acid (aa) identity with three other R-Spondin family members (1, 2). All R-Spondins regulate Wnt/ beta-Catenin signaling but have distinct expression patterns (1-3). R-Spondin 1 competes with the Wnt antagonist DKK-1 for binding to the Wnt co-receptors, Kremen and LRP-6, reducing their DKK-1-mediated internalization (4). However, reports are mixed on whether R-Spondin 1 binds LRP-6 directly (4-6). R-Spondin 1 is expressed in early development at the roof plate boundary and is thought to contribute to dorsal neural tube development (3, 7). In humans, rare disruptions of the R-Spondin 1 gene are associated with tendencies for XX sex reversal (phenotypic male) or hermaphroditism, indicating a role for R-Spondin 1 in gender-specific differentiation (7, 8). Mutations in R-Spondin 1 are also linked with palmoplantar keratoderma, abnormal thickening of the skin on the palms of the hands and soles of the feet (7, 8). Postnatally, R-Spondin 1 is expressed by neuroendocrine cells in the intestine, adrenal gland and pancreas, and by epithelia in kidney and prostate (9). Injection of recombinant R-Spondin 1 in mice causes activation of beta-catenin and proliferation of intestinal crypt epithelial cells, and ameliorates experimental colitis (9, 10). Interest in R-Spondin 1 as a cell culture supplement has grown with the expansion of the organoid field. R-Spondin 1 is widely used in organoid cell culture workflows as a vital component that promotes both growth and survival of 3D organoids (11).

Structurally similar to other R-Spondins, R-Spondin 1 contains two adjacent cysteine-rich furin-like domains (aa 34-135) with one potential N-glycosylation site, followed by a thrombospondin (TSP-1) motif (aa 147-207) and a region rich in basic residues (aa 211-263). Only the furin-like domains are needed for beta-catenin stabilization (2, 12). A putative nuclear localization signal at the C-terminus may allow some expression in the nucleus (13). Potential isoforms of 200 and 236 aa have an alternate, shorter N-terminus or are missing aa 146-208, respectively (14). Over aa 21-263, human R-Spondin 1 shares 89%, 87%, 92%, 91%, 91% and 89% aa identity with mouse, rat, horse, dog, goat, and cow RSPO-1, respectively.

References
  1. Chen, J-Z. et al. (2002) Mol. Biol. Rep. 29:287.
  2. Kim, K.-A. et al. (2006) Cell Cycle 5:23.
  3. Nam, J.-S. et al. (2007) Gene Expr. Patterns 7:306.
  4. Binnerts, M.E. et al. (2007) Proc. Natl. Acad. Sci. USA 104:14700.
  5. Nam, J.-S. et al. (2006) J. Biol. Chem. 281:13247.
  6. Wei, Q. et al. (2007) J. Biol. Chem. 282:15903.
  7. Kamata, T. et al. (2004) Biochim. Biophys. Acta 1676:51.
  8. Parma, P. et al. (2006) Nat. Genet. 38:1304.
  9. Kim, K.-A. et al. (2005) Science 309:1256.
  10. Zhao, J. et al. (2007) Gastroenterology 132:1331.
  11. Drost and Clevers. (2018) Nature Reviews Cancer 18:407.
  12. Kazanskaya, O. et al. (2004) Dev. Cell 7:525.
  13. Tomaselli, S. et al. (2008) Hum. Mutat. 29:220.
  14. UniProt # Q2MKA7.
Long Name
Roof Plate-specific Spondin 1
Entrez Gene IDs
284654 (Human); 192199 (Mouse); 102122369 (Cynomolgus Monkey)
Alternate Names
Cristin 3; CRISTIN3; FLJ40906Roof plate-specific spondin-1; HRspo1; roof plate-specific spondin; RSPO; RSPO1; RSpondin 1; R-Spondin 1; R-spondin homolog (Xenopus laevis); RSPONDIN; R-spondin1; R-spondin-1

Product Specific Notices

The above product was manufactured, tested and released by R&D System's contract manufacturer, Qkine Ltd, at 1 Murdoch House, Cambridge, UK, CB5 8HW. The product is for research use only and not for the diagnostic or theraputic use.

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