Meet the newest class of Amphipols. They're called PMAL.
And they'll help you set new standards in membrane protein research.

First described in 1996 by Jean-Luc Popot, amphipols are a class of polymers that can stabilize membrane proteins in a detergent-free, aqueous solution(1). After the solubilization and purification of a membrane protein in detergent, amphipols can replace the detergent, and excess detergent can be removed from the solution. The most well characterized amphipol is Amphipol A8-35, with many biophysical applications, including structure determination via single-particle electron cryo-microscopy (Cryo-EM)(2, 3). To date, there have been over 20 Cryo-EM structures of membrane proteins determined using amphipols, the majority using Amphipol A8-35.


In recent years, another class of amphipols, our PMAL series, has been gaining traction for use in Cryo-EM, with a number of structures published in the past two years. Developed in a collaboration between Charles Sanders (Vanderbilt University) and Anatrace(4, 5), PMAL amphipols are zwitterionic, and contain repeating units of a carboxyl, ammoniumamidate, and alkyl chain (C8, C12, or C16). This is in contrast to Amphipol A8-35, which is negatively charged and comprises octylamine and isopropylamine groups randomly grafted on a polyacrylate chain. To date, for Cryo-EM studies, the PMAL-C8 molecule has been the most successful of the three versions. Some of the recent Cryo-EM structures using PMAL-C8 include:

  • April 2015: The labs of Yifan Cheng and David Julius at UCSF determined the structure of the TRP ion channel TRPA1 (PDB: 3J9P)(6). This protein detects noxious compounds that elicit pain and cause neurogenic inflammation. TRPA1 was solubilized and purified using LMNG, and reconstituted into Amphipol PMAL-C8 (1:3 ratio). Prior to structure determination, the excess detergent was removed by Bio Beads.
  • July 2017: The complex of the ERAD channel Hrd1 in complex with Hrd3 was determined by the labs of Tom Rapoport and Maofu Liao at Harvard University (PDB: 5V6P)(7). The Hrd1 dimer forms a channel for the movement of misfolded polypeptides through the endoplasmic reticulum membrane. This complex was solubilized and purified in DM, and reconstituted into Amphipol PMAL-C8 (1:3 ratio) for Cryo-EM. The excess detergent was removed by dilution and concentration of the sample.
  • December 2017: The labs of Jue Chen and Roderick MacKinnon at Rockefeller University used Amphipol PMAL-C8 in the structure determination of the Human pancreatic KATP channel (6C3O and 6C3P)(8). Comprising the ABC transporter SUR1, and the K+ channel Kir6.2, KATP channels respond to intracellular levels of ATP and ADP to regulate the control of a number of cellular processes, including the production of insulin. For solubilization, LMNG:CHS was utilized, and the complex was purified in Digitonin and a 3:1:1 mix of the phospholipids POPC, POPE, and POPG. The channel was exchanged into Amphipol PMAL-C8 (1:10 ratio) for structure determination, and the detergent removed by dilution and concentration.
  • March 2018: In the most recent example, the SLC4A4 sodium-coupled acid-base transporter NBCe1 was determined by the labs of Z. Hong Zhou and Ira Kurtz at UCLA (PDB: 6CAA)(9). Mutations of this transporter cause a number of deficiencies, including blindness, migraines, and impaired cognitive function. To prepare the NBCe1 protein for Cryo-EM studies, solubilization was performed using Triton X-100, and purification using DDM. The protein was exchanged into Amphipol PMAL-C8 (1:3 ratio), and detergent removed using Bio-Beads.

A New Book on Amphipols:
We are excited to announce the publication of a new book by Jean-Luc Popot, “Membrane Proteins in Aqueous Solutions: From Detergents to Amphipols”. This book examines the many ways in which membrane proteins can be handled outside of their native lipid environment. Topics include a thorough overview of many of the technologies developed over the past twenty years, including: detergents, bicelles, nanodiscs, amphipathic peptides, fluorinated surfactants, and a particular emphasis on amphipols.

 

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