Expression of a highly antigenic and native-like folded extracellular domain of the human α1 subunit of muscle nicotinic acetylcholine receptor, suitable for use in antigen specific therapies for Myasthenia Gravis

Athanasios Niarchos; Marios Zouridakis; Vassilis Douris; Assimina Georgostathi; Dimitra Kalamida; Alexandros Sotiriadis; Konstantinos Poulas; Kostas Iatrou; Socrates J Tzartos

doi:10.1371/journal.pone.0084791

Expression of a highly antigenic and native-like folded extracellular domain of the human α1 subunit of muscle nicotinic acetylcholine receptor, suitable for use in antigen specific therapies for Myasthenia Gravis

PLoS One. 2013 Dec 20;8(12):e84791. doi: 10.1371/journal.pone.0084791. eCollection 2013.

Authors

Athanasios Niarchos¹, Marios Zouridakis², Vassilis Douris³, Assimina Georgostathi¹, Dimitra Kalamida¹, Alexandros Sotiriadis², Konstantinos Poulas¹, Kostas Iatrou³, Socrates J Tzartos⁴

Affiliations

¹ Department of Pharmacy, University of Patras, Patras, Greece.
² Department of Biochemistry, Hellenic Pasteur Institute, Athens, Greece.
³ Institute of Biosciences and Applications, National Centre for Scientific Research "Demokritos", Athens, Greece.
⁴ Department of Pharmacy, University of Patras, Patras, Greece ; Department of Biochemistry, Hellenic Pasteur Institute, Athens, Greece.

Abstract

We describe the expression of the extracellular domain of the human α1 nicotinic acetylcholine receptor (nAChR) in lepidopteran insect cells (i-α1-ECD) and its suitability for use in antigen-specific therapies for Myasthenia Gravis (MG). Compared to the previously expressed protein in P. pastoris (y-α1-ECD), i-α1-ECD had a 2-fold increased expression yield, bound anti-nAChR monoclonal antibodies and autoantibodies from MG patients two to several-fold more efficiently and resulted in a secondary structure closer to that of the crystal structure of mouse α1-ECD. Our results indicate that i-α1-ECD is an improved protein for use in antigen-specific MG therapeutic strategies.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Autoantibodies / blood
Autoantibodies / metabolism*
Blotting, Western
Bungarotoxins / metabolism
Cell Line
Circular Dichroism
Cloning, Molecular
Electrophoresis, Polyacrylamide Gel
Enzyme-Linked Immunosorbent Assay
Epitopes
Humans
Immunosorbent Techniques
Iodine Radioisotopes / metabolism
Moths
Muscle, Skeletal / metabolism*
Myasthenia Gravis / immunology*
Myasthenia Gravis / therapy*
Pichia
Plasmids / genetics
Protein Structure, Tertiary / physiology
Radioimmunoassay
Receptors, Nicotinic / metabolism*
Receptors, Nicotinic / therapeutic use*

Substances

Autoantibodies
Bungarotoxins
CHRNA1 protein, human
Epitopes
Iodine Radioisotopes
Receptors, Nicotinic

Grants and funding

This work was supported by grants from the European Commission (FP7 FightMG no. 242210, and FP7 REGPOT 2010-1 NeuroSign no. 264083) and the Greek NSRF “Thalis” Autoimmunity grant. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.