Recombinant Human Sonic Hedgehog/Shh (C24II) Animal-Free, CF

Animal-Free™ Recombinant Human Sonic Hedgehog/Shh (Catalog # AFL1845) induces alkaline phosphatase production by the C3H10T1/2 mouse embryonic fibroblast cell line. The ED50 for this effect is 0.1-0.4 μg/mL.

1 μg/lane of Animal-Free™ Recombinant Human Sonic Hedgehog/Shh (C24II) Protein (Catalog # AFL1845) was resolved with SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by silver staining, showing bands at 22 kDa.

• Reactivity: Hu
• Applications: Bioactivity
• Format: Carrier-Free

Recombinant Human Sonic Hedgehog/Shh (C24II) Animal-Free, CF Summary

• Additional Information: Intended for preclinical researchers who may transition to GMP Sonic Hedgehog/Shh (C24II) for their clinical work
• Details of Functionality: Measured by its ability to induce alkaline phosphatase production by C3H10T1/2 mouse embryonic fibroblast cells. Nakamura, T. et al. (1997) Biochem. Biophys. Res. Commun. 237:465. The ED₅₀ for this effect is 0.1-0.4 µg/mL.
• Source: E. coli-derived human Sonic Hedgehog/Shh protein
  Cys24-Gly197 (Cys24Ile-Ile), with an N-terminal Met
  Produced using non-animal reagents in an animal-free laboratory.
• Accession #: NP_000184.1
• N-terminal Sequence: Met
• Protein/Peptide Type: Animal-Free Recombinant Proteins
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
• Endotoxin Note: <0.01 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Dilutions:
  • Bioactivity
• Theoretical MW: 20 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• SDS-PAGE: 22 kDa, reducing conditions

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 3 months, -20 to -70 °C under sterile conditions after reconstitution.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS and NaCl.
• Purity: >95%, by SDS-PAGE under reducing conditions and visualized by silver stain.
• Reconstitution Instructions: Reconstitute at 100-200 μg/mL in PBS, and allow up to 24 hours at 2 to 8 °C for complete reconstitution.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human Sonic Hedgehog/Shh (C24II) Animal-Free, CF

• HHG1
• HHG-1
• HLP3
• HPE3
• MCOPCB5
• MCOPCB5sonic hedgehog (Drosophila) homolog
• Shh
• ShhNC
• SMMCI
• SMMCIsonic hedgehog homolog (Drosophila)
• sonic hedgehog homolog
• sonic hedgehog protein
• Sonic Hedgehog
• TPT
• TPTPS

Background

Sonic Hedgehog (Shh) is expressed in embryonic tissues that are critical for the patterning of the developing central nervous system, somite, and limb. It is also involved in whisker, hair, foregut, tooth, and bone development. Shh regulates neural and hematopoietic stem cell fate and is important for thymocyte differentiation and proliferation as well as T cell determination. In adult tissue Shh is associated with cancer development and tissue remodeling following injury (1-3). Human Shh encodes a 462 amino acid (aa) precursor protein that is autocatalytically processed to yield a non-glycosylated 19 kDa N-terminal fragment (Shh-N) and a glycosylated 25 kDa C-terminal protein (Shh-C) (4). Shh-C, which is responsible for the intramolecular processing of Shh, is rapidly degraded following Shh proteolysis (5). Shh-N is highly conserved, sharing >98% aa identity between mouse, human, rat, canine, porcine, and chicken Shh-N. Shh-N can be palmitoylated at its N-terminal cysteine and modified by cholesterol addition at its C-terminus (6). These modifications contribute to the membrane tethering of Shh as well as its assembly into various sized multimers (6-9). Lipid modification and multimerization greatly increase Shh-N receptor binding affinity and signaling potency (5, 6, 8, 9). Monomeric and multimeric Shh can be released from the plasma membrane by the cooperative action of DISP1, SCUBE2, and TACE/ADAM17 (10-12). Modifications also extend the effective range of Shh functionality and are required for the development of protein gradients important in tissue morphogenesis (9, 13). Canonical signaling of Shh is mediated by a multicomponent receptor complex that includes Patched (PTCH1, PTCH2) and Smoothened (SMO) (14). The binding of Shh to PTCH releases the basal repression of SMO by PTCH. Shh activity can also be regulated through interactions with heparin, glypicans, and membrane-associated Hip (hedgehog interacting protein) (13, 15, 16).

1. Briscoe, J. and P.P. Therond (2013) Mol. Cell. Biol. 14:416.
2. Aviles, E.C. et al. (2013) Front. Cell. Neurosci. 7:86.
3. Xie, J. et al. (2013) OncoTargets Ther. 6:1425.
4. Marigo, V. et al. (1995) Genomics 28:44.
5. Zeng, X. et al. (2001) Nature 411:716.
6. Feng, J. et al. (2004) Development 131:4357.
7. Goetz, J.A. et al. (2006) J. Biol. Chem. 281:4087.
8. Pepinsky, R.B. et al. (1998) J. Biol. Chem. 273:14037.
9. Chen, M.-H. et al. (2004) Genes Dev. 18:641.
10. Etheridge, L.A. et al. (2010) Development 137:133.
11. Jakobs, P. et al. (2014) J. Cell Sci. 127:1726.
12. Dierker, T. et al. (2009) J. Biol. Chem. 284:8013.
13. Lewis, P.M. et al. (2001) Cell 105:599.
14. Carpenter, D. et al. (1998) Proc. Natl. Acad. Sci. USA 95:13630.
15. Filmus, J. and M. Capurro (2014) Matrix Biol. 35:248.
16. Chuang, P.-T. and A.P. McMahon (1999) Nature 397:617.

Bioinformatics

• Uniprot:
  • NP_000184.1()