Structure and function of myelin proteins

 

Myelin is an ingenious specialization of the nervous system, enabling efficient, fast transmission along neural fibers of moderate diameters. By virtue of its extraordinary high lipid content, the myelin membrane stack around axons can act as an electrical insulator. Albeit outnumbered by lipids, the bulk of the myelin proteome consists of highly specialized membrane proteins that are indispensable for the proper assembly and maintenance of the myelin sheath, as alterations in abundance or structure of these proteins are the cause of many neuropathies. Surprisingly, the molecular mechanisms of most myelin proteins are not very well explored so far. David Ewers has a keen interest in proteins that interact with membranes, and his research is focused on molecular mechanisms of myelin proteins.

Predicted 3D structure of PMP22 with known and putative functional residues highlighted (Jumper et al. Highly accurate protein structure prediction with AlphaFold (Nature 2021)).
 

PMP22

The peripheral myelin protein of 22 kDa (PMP22) is an integral membrane protein with high abundance in the myelin sheaths of peripheral nerves. Overexpression of PMP22 in myelin-forming Schwann cells causes the most frequent hereditary peripheral neuropathy, Charcot-Marie-Tooth disease 1A (CMT1A). Despite more than three decades of research on this important disease-causing protein, it is still unclear what role PMP22 plays in the physiology of Schwann cells. We try to elucidate this role as well as more general functions of PMP22, which is expressed in many cell types throughout the body albeit at much lower levels than in Schwann cells. Using co-immunoprecipitation, affinity pull-down and proximity ligation assays, we currently focus on interaction partners (other proteins, lipids) that link PMP22 to important growth signaling cascades. Furthermore, we express PMP22 at different scales in a variety of mammalian cells to analyze its subcellular location as well as its impact on cellular signaling. We also purify the protein from mammalian cells for in-depth characterization of quarternary structure and interactions of PMP22 in absence or selected presence of other cellular factors.

HEK cells expressing membrane anchored GFP (green) and PMP22-mCherry (red).
 

Available methods for myelin protein research

  • A wide range of liquid chromatography methods, including
    • Fluorescent detection size exclusion chromatography (FSEC)
    • SEC coupled to Right angle light scattering (SEC-RALS)
    • Affinity chromatography
    • High pressure liquid chromatography (HPLC)
  • Fluorescence spectroscopy
  • Differential scanning fluorimetry (DSF)
  • Fluorescent thermal shift assay (TSA)
  • Isothermal titration calorimetry
  • Single molecule transmission electron microscopy (TEM)
A single oligomer species of ALFA-tagged PMP22 heterologously expressed in HEK cells as shown by FSEC. Elution of DDM-solubilized, ALFA-tagged PMP22 was detected via a fluorescent Anti-ALFA single-domain antibody. 

A single oligomer species of ALFA-tagged PMP22 heterologously expressed in HEK cells as shown by FSEC. Elution of DDM-solubilized, ALFA-tagged PMP22 was detected via a fluorescent Anti-ALFA single-domain antibody.
 

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