Amphipol-Assisted Folding of Membrane
Proteins
6
Summary
Folding membrane proteins (MPs) in vitro is important from both practical and
theoretical points of view. Amphipols (APols) have emerged as an interesting medium
in which to fold MPs. Both α-helical and β-barrel MPs are amenable to folding in
APols, using simple procedures and achieving high yields. All APols hitherto tested
(A8-35, sulfonated APols, non-ionic APols) have proven to provide a medium favorable
to MP folding. From a practical point of view, the approach is particularly interesting
when applied to MPs that have been produced in large amounts in inclusion bodies,
from which they can be recovered only under a denatured state. The resulting MP/APol
complexes can be used as such or the protein transferred to other media more
appropriate to a given type of experiment (e.g. to lipidic mesophases for crystallization). From a theoretical point of view, the observation that MPs can fold very
efficiently in a medium so different from a lipid bilayer raises interesting questions
about the forces that determine the final fold.
6.1
Introduction
Folding a denatured or unfolded membrane protein (MP) to a functional form is one of the toughest
challenges of membrane biochemistry (for reviews, see e.g. Booth 2003; Kiefer 2003; White 2003;
Tamm et al. 2004; Bowie 2005; Bannwarth and Schulz 2003; Michaux et al. 2008; Stanley and
Fleming 2008; Harris and Booth 2012; Otzen and Andersen 2013; Popot 2014; Kleinschmidt 2015). It
is considered by many as akin to reaching an isolated spot of solid land in the middle of a treacherous
swamp, spotted with potholes and quicksand, with no paths marked. Many a laboratory director
hesitates before committing students or postdoctoral fellows to such a project, for fear of ruining
their career. Yet, most MPs are too rare to be purified from natural sources in amounts sufficient for
structural and extensive functional studies. Overexpressing them in large amounts under a functional
form is tricky, costly, lengthy to set up and optimize, and, more often than not, frustrating. This is
because targeting an overexpressed MP to a membrane, where it can insert and achieve its native
structure, tends to be toxic, resulting in limited expression yields, and only a fraction of the proteins
# Springer International Publishing AG, part of Springer Nature 2018
J. -L. Popot, Membrane Proteins in Aqueous Solutions, Biological and Medical Physics,
Biomedical Engineering, https://doi.org/10.1007/978-3-319-73148-3_6
333
Proteins
6
Summary
Folding membrane proteins (MPs) in vitro is important from both practical and
theoretical points of view. Amphipols (APols) have emerged as an interesting medium
in which to fold MPs. Both α-helical and β-barrel MPs are amenable to folding in
APols, using simple procedures and achieving high yields. All APols hitherto tested
(A8-35, sulfonated APols, non-ionic APols) have proven to provide a medium favorable
to MP folding. From a practical point of view, the approach is particularly interesting
when applied to MPs that have been produced in large amounts in inclusion bodies,
from which they can be recovered only under a denatured state. The resulting MP/APol
complexes can be used as such or the protein transferred to other media more
appropriate to a given type of experiment (e.g. to lipidic mesophases for crystallization). From a theoretical point of view, the observation that MPs can fold very
efficiently in a medium so different from a lipid bilayer raises interesting questions
about the forces that determine the final fold.
6.1
Introduction
Folding a denatured or unfolded membrane protein (MP) to a functional form is one of the toughest
challenges of membrane biochemistry (for reviews, see e.g. Booth 2003; Kiefer 2003; White 2003;
Tamm et al. 2004; Bowie 2005; Bannwarth and Schulz 2003; Michaux et al. 2008; Stanley and
Fleming 2008; Harris and Booth 2012; Otzen and Andersen 2013; Popot 2014; Kleinschmidt 2015). It
is considered by many as akin to reaching an isolated spot of solid land in the middle of a treacherous
swamp, spotted with potholes and quicksand, with no paths marked. Many a laboratory director
hesitates before committing students or postdoctoral fellows to such a project, for fear of ruining
their career. Yet, most MPs are too rare to be purified from natural sources in amounts sufficient for
structural and extensive functional studies. Overexpressing them in large amounts under a functional
form is tricky, costly, lengthy to set up and optimize, and, more often than not, frustrating. This is
because targeting an overexpressed MP to a membrane, where it can insert and achieve its native
structure, tends to be toxic, resulting in limited expression yields, and only a fraction of the proteins
# Springer International Publishing AG, part of Springer Nature 2018
J. -L. Popot, Membrane Proteins in Aqueous Solutions, Biological and Medical Physics,
Biomedical Engineering, https://doi.org/10.1007/978-3-319-73148-3_6
333