Exploring the Interplay of Lipids and Membrane proteins
Doctoral thesis, 2014
The interplay between lipids and membrane proteins is known to affect
membrane protein topology and thus have significant effect (control) on
their functions. In this PhD thesis, the influence of lipids on the membrane
protein function was studied using three different membrane protein models.
A monotopic membrane protein, monoglucosyldiacylglyecerol synthase
(MGS) from Acholeplasma laidlawii is known to induce intracellular vesicles
when expressed in Escherichia coli. The mechanism leading to this unusual
phenomenon was investigated by various biochemical and biophysical
techniques. The results indicated a doubling of lipid synthesis in the cell,
which was triggered by the selective binding of MGS to anionic lipids. Multivariate
data analysis revealed a good correlation with MGS production.
Furthermore, preferential anionic lipid sequestering by MGS was shown to
induce a different fatty acid modeling of E. coli membranes. The roles of
specific lipid binding and the probable mechanism leading to intracellular
vesicle formation were also investigated.
As a second model, a MGS homolog from Synechocystis sp. PCC6803 was
selected. MgdA is an integral membrane protein with multiple transmembrane
helices and a unique membrane topology. The influence of different
type of lipids on MgdA activity was tested with different membrane fractions
of Synechocystis. Results indicated a very distinct profile compared to
Acholeplasma laidlawii MGS. SQDG, an anionic lipid was found to be the
species of the membrane that increased the MgdA activity 7-fold whereas
two other lipids (PG and PE) had only minor effects on MgdA. Additionally,
a working model of MgdA for the biosynthesis and flow of sugar lipids between
Synechocystis membranes was proposed.
The last model system was another integral membrane protein with a distinct
structure but also a different function. The envelope stress sensor, CpxA and
its interaction with E. coli membranes were studied. CpxA autophosphorylation
activity was found to be positively regulated by phosphatidylethanolamine
and negatively by anionic lipids. In contrast, phosphorylation of CpxR
by CpxA revealed to be increased with PG but inhibited by CL. Non-bilayer
lipids had a negative impact on CpxA phosphotransfer activity.
Taken together, these studies provide a better understanding of the significance
of the interplay of lipids and model membrane proteins discussed here.