Biotinylated Amphipol A8-35:
Raising the standards of your binding studies.

In recent years, amphipols have been increasingly used for the study of membrane proteins(1, 2). These polymers are able to stabilize membrane proteins in an aqueous environment in the absence of detergent. The most thoroughly characterized amphipol, A8-35, contains a short ~35 residue hydrophilic polyacrylate chain with randomly grafted octylamine (25%) and isopropylamine (40%) groups. Among other applications, Amphipol A8-35 has become an invaluable tool in the Cryo-EM studies of membrane protein. Some of the Cryo-EM structures of membrane proteins determined using Amphipol A8-35 include: γ-secretase (PDB: 5A63)(3), TRPV2 (PDB: 5AN8)(4, 5), STRA6 (PDB: 5SY1)(6), Bovine Respirasome (PDB: 5LUF)(7), the Holo-translocon (PDB: 5MG3)(8), Polycystin-2 (PDB: 5MKE)(9), 1and the Cyclic-nucleotide gated channel (PDB: 5H3O)(10). To further expand their applications to membrane proteins, there is continuous work in the functionalization of amphipols with various labels, dyes, and tags(11, 12).

Anatrace is excited to offer a new, biotinylated A8-35 which allows for the immobilization of membrane proteins onto solid supports. First described in 2009 by the lab of Jean-Luc Popot(13), membrane proteins reconstituted into biotinylated A8-35 can be immobilized onto surfaces coated with streptavidin. To demonstrate the application of biotinylated A8-35 to surface plasmon resonance (SPR) assays, four different membrane proteins were reconstituted into biotinylated A8-35, immobilized onto an streptavidin coated SPR chip, and binding to protein-specific antibodies were measured. To show that ligand binding could be measured, the nicotinic acetylcholine receptor (nAChR) was reconstituted into biotinylated A8-35 and bound to streptavidin coated beads. When fluorescently labeled ligand was added, a fluorescent signal was monitored at the surface of the beads by confocal fluorescence microscopy, indicating an interaction of the ligand with the agonist-binding site of nAChR. In another study, biotinylated A8-35 has been shown to be compatible with have been recently shown to be compatible with nanoscale electrodes for functional membrane protein arrays(14)

Figure 1: New Biotinylated Amphipol A8-35 (BAM01)



 

Upcoming Meetings
Anatrace is excited to sponsor the upcoming Mid-Atlantic Molecular Crystallography Meeting in Baltimore, MD. This meeting is hosted by The University of Maryland School of Medicine. Anatrace will also be attending the 24th Congress of the International Union of Crystallography Meeting in Hydrabad, India from August 21-28. Stay up to date on all of the conferences Anatrace will be attending by visiting our Events Calendar.


 
References:
  1. Zoonens, M. and Popot, J. L. (2014) J Membr Biol., 247(9-10), 759-796.
  2. Popot, J. L., et al. (2003) Cell Mol Life Sci., 60(8), 1559-1574.
  3. Bai, X. C., et al. (2015) Nature, 525(7568), 212-217.
  4. Zubcevic, L. et al. (2016) Nat Struct Mol Biol., 23(2), 180-186.
  5. Huynh, K. W., et al. (2014) J Membr Biol., 247(9-10), 843-851.
  6. Chen, Y., et al. (2016) Science, 353(6302), pii: aad8266. doi: 10.1126/science.aad8266.
  7. Sousa, J. S., et al. (2016), Elife, 10(5), pii: e21290. doi: 10.7554/eLife.21290.
  8. Botte, M., et al. (2016) Sci. Rep. 6(38399), doi: 10.1038/srep38399.
  9. Wilkes, M., et al. (2017) Nat Struct Mol Biol., 24(2), 123-130.
  10. Li, M., et al. (2017) Nature, 542(7639), 60-65.
  11. Le Bon, C., et al. (2014) J Membr Biol., 247(9-10), 797-814.
  12. Della Pia, E. A., et al. (2014) J Membr Biol., 247(9-10), 815-826.
  13. Charvolin, D., et al. (2009) Proc Natl Acad Sci U S A, 106(2), 405-410.
  14. Della Pia, E. A., et al. (2014) ACS Nano., 8(2), 1844-1853.