Staff Members of the Institute of Biochemistry, TU - Institut für ...

investigated the effect of squalene on biophysical properties of lipid particles and membranes
and compared these results to artificial membranes. Our experiments showed that squalene
lowered the order of SE which form concentric shells in lipid droplets from wild type. In
biological and artificial membranes fluorescence spectroscopy studies revealed that changes
in membrane fluidity/rigidity are not result of absolute squalene levels, but are rather affected
by the squalene to ergosterol ratio. In a fluid membrane environment squalene induces rigidity
of the membrane, whereas in rigid membrane there is almost no additive effect of squalene.
Although squalene is not a typical membrane lipid it may be regarded as a mild modulator of
biophysical membrane properties.
Pichia pastoris organelles and lipids
Doctoral Thesis completed
Miroslava Spanova: Neutral Lipid Storage in Yeast
The yeast Pichia pastoris is an important experimental
system for heterologous expression of proteins.
Nevertheless, surprisingly little is known about organelles of
this microorganism. For this reason, we started a systematic
biochemical and cell biological study to establish
standardized methods of Pichia pastoris organelle isolation
and characterization. Recent work focused on the
biochemical characterization of the plasma membrane and
secretory organelles from Pichia pastoris. Moreover, lipid
droplets from Pichia pastoris were isolated and analyzed
with respect to their lipids and proteins. Mutants bearing
defects in non-polar lipid formation of Pichia pastoris were
constructed. Detailed investigations with these strains are
currently in progress. Methods of Pichia pastoris organelle
characterization include standardized techniques of lipidome
and proteome analyses.
Transmission electron micrographs of Pichia pastoris wild
type cells grown on glucose (A), glycerol (B), and methanol
(C). Pictures shown be courtesy of Dr. G. Zellnig, University
of Graz, Austria
Squalene, a natural triterpene, is a key intermediate in ergosterol synthesis of the yeast and
present in the cell only at minor amounts under standard conditions. For the present study we
constructed strains of Saccharomyces cerevisiae accumulating squalene to perform
fundamental research but also as a possible source for biotechnological production of this
lipid. We first addressed localization of squalene within the cell and its possible lipotoxic
effect in the yeast. We showed that the highly hydrophobic squalene preferentially localizes to
lipid particles/droplets. In this compartment it decreases the order which is created by steryl
esters forming concentric layers around a core of triacylglycerols. We also observed that
squalene did not exhibit a lipotoxic effect even in mutant strains which are unable to form
lipid particles. In strains devoid of lipid particles large amounts of squalene were stored in
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