Biology Reference
In-Depth Information
We analyzed individually 7700 vesicles in the case of ferritin entrapment, and 400 vesicles
in the case of ribosome entrapment (the third case, namely the entrapment of the PURE
system, was not analyzed quantitatively but only qualitatively). As shown in
Figure 14.2C
,
samples consisted of many empty vesicles and very few solute
vesicles. The
classical Poisson model cannot explain the existence of such vesicles. The experimentally
determined solute occupancy distribution, in all tested cases, does not follow the Poisson
distribution, but it is rather shaped like
power-law
distributions. In particular, a long tail of
decreasing probabilities (0.1
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super-filled
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1%) was typically found in all the experimentally determined
distributions, indicating that the probability of finding vesicles containing a very high
number of solutes (ferritins, or ribosomes) was low, but not so unrealistic as estimated by
the Poisson curve.
Considering the experimentally determined number of macromolecules inside
liposomes of a certain diameter, it is possible to determine the internal concentration of
these macromolecules in each liposome. The results indicate that for
vesicles,
the internal concentration can be up to ca. 20 times higher (but typical values are around
12 for ferritin and around 6 for ribosomes) than the external bulk solute concentration
(exceptionally high internal concentrations have been also recorded in rare cases, up to
about 50 times the expected concentration). This effect is size-dependent: smaller vesicles
(diameter
superfilled
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'
,
100 nm) reach higher enhancement factors when compared to larger vesicles.
Having demonstrated that the spontaneous formation of vesicles (in a solute-containing
solution) brings about not only empty and regularly filled vesicles, but also a small but
measurable fraction of
vesicles, we have speculated about the possible generative
mechanisms. Our working hypothesis, at the moment, is based on the interplay between
solute/bilayer interaction and the mechanism of vesicle formation.
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Let us discuss the vesicle
closure mechanism that is thought to be involved in several vesicle formation processes such
as the injection method. Open bilayer fragments, formed by self-association of lipids, are the
intermediates that give rise to closed vesicles. Their half-life has been estimated in the order of
milliseconds. We suppose that the solutes stochastically adsorb on the bilayer fragments and
slow down the closure rate so that additional solutes can be recruited by the still-open
bilayer. The expulsion of bound water would be the driving force of the process.
super-filled
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A possible biological relevance of these results and their importance in the origin-of-life
scenarios can be discussed as follows. The spontaneous formation of first membrane-
enclosed compartments might have been fundamental to the origins of primitive cells.
These results suggest that the compartment is no longer a simple physical container, but it
becomes an agent that induces reactivity under conditions where reactions are not possible
(diluted solutions). If we assume the possible development of macromolecules and some
metabolic network outside compartments, the fact that a large number of solutes can indeed
be unexpectedly entrapped inside lipid vesicles during their formation would provide good
evidence to explain the emergence of functional primitive cells from free components
(separated solutes and lipids). In fact, one of the major theoretical and practical obstacles to
explain the formation of functional primitive cells is the very limited solute permeability of
most membranes. The other interesting consequence of the
of solutes
inside vesicles is that the resulting high concentration of enzymes or macromolecules allows
the overcoming of critical concentration for effective reactivity (e.g. overcoming the
dissociation constant of molecular complexes, or enhancing reaction rates of enzymatic
reaction when the substrate concentration is higher than the Michaelis constant). This effect,
therefore, means that the intravesicle water pool can be considered a privileged environment
for biochemical transformation. Clearly, this is also linked to the concept of cellular
crowding. The results suggest that a kind of internal crowding can spontaneously emerge
from unexpected physical events, probably triggered by the interplay of stochastic
fluctuations and the mechanism of vesicle formation.
super-concentration
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