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Physical Routes to Primitive Cells: An Experimental Model Based on the Spontaneous Entrapment of Enzymes inside Micrometer-Sized Liposomes.

D'Aguanno E, Altamura E, Mavelli F, Fahr A, Stano P, Luisi PL - Life (Basel) (2015)

Bottom Line: We have reported that proteins and ribosomes can be encapsulated very efficiently, against statistical expectations, inside a small number of liposomes.Moreover the transcription-translation mixture, which realistically mimics a sort of minimal metabolic network, can be functionally reconstituted in liposomes owing to a self-concentration mechanism.Here we firstly summarize the recent advancements in this research line, highlighting how these results open a new vista on the phenomena that could have been important for the formation of functional primitive cells.

View Article: PubMed Central - PubMed

Affiliation: Science Department, Roma Tre University, Viale G. Marconi 446, I-00146 Rome, Italy. Alfred.Fahr@uni-jena.de.

ABSTRACT
How did primitive living cells originate? The formation of early cells, which were probably solute-filled vesicles capable of performing a rudimentary metabolism (and possibly self-reproduction), is still one of the big unsolved questions in origin of life. We have recently used lipid vesicles (liposomes) as primitive cell models, aiming at the study of the physical mechanisms for macromolecules encapsulation. We have reported that proteins and ribosomes can be encapsulated very efficiently, against statistical expectations, inside a small number of liposomes. Moreover the transcription-translation mixture, which realistically mimics a sort of minimal metabolic network, can be functionally reconstituted in liposomes owing to a self-concentration mechanism. Here we firstly summarize the recent advancements in this research line, highlighting how these results open a new vista on the phenomena that could have been important for the formation of functional primitive cells. Then, we present new evidences on the non-random entrapment of macromolecules (proteins, dextrans) in phospholipid vesicle, and in particular we show how enzymatic reactions can be accelerated because of the enhancement of their concentration inside liposomes.

No MeSH data available.


Related in: MedlinePlus

(a) Ferritin occupancy distribution as determined by analyzing about 7700 individual vesicles imaged by cryo-TEM. Experimental points shown as colored circles, theoretical expectations as empty circles connected by a line; (b) Two conventional POPC vesicles: one is super-filled with ferritin (encapsulating much more ferritin molecules than expected), the other does not contain any ferritin molecule (also against expectations). Images reproduced from [10] with the permission of Wiley.
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life-05-00969-f004: (a) Ferritin occupancy distribution as determined by analyzing about 7700 individual vesicles imaged by cryo-TEM. Experimental points shown as colored circles, theoretical expectations as empty circles connected by a line; (b) Two conventional POPC vesicles: one is super-filled with ferritin (encapsulating much more ferritin molecules than expected), the other does not contain any ferritin molecule (also against expectations). Images reproduced from [10] with the permission of Wiley.

Mentions: Intrigued by these unexpected conclusions, we then started a direct investigation on solute encapsulation inside spontaneously formed lipid vesicles. We recalled our previous work on the use of the protein ferritin as a marker for the vesicle lumen [4,5]. Individual ferritin molecules contain high amount of iron in form of hydrous ferric oxide phosphate, and it is widely used in electronmicroscopy. Since the number of ferritin molecules inside vesicles can be directly counted in images obtained by cryo-transmission electronmicroscopy (cryo-TEM), and the vesicle size is measurable as well, it is possible to measure the solute occupancy distribution f(n) in vesicle populations and compare it with the expected Poisson distribution, p(n). As shown in Figure 4A, the observed distribution is quite different than the theoretical one. The distribution is not bell-shaped, and—at high n—the measured distribution has a long “tail” of values that are significantly higher than the vanishing small values of the Poisson curve. The experimentally determined distribution strongly resemble a power law, i.e., f(n) ~ 1/na (a > 0), and Figure 4B summarizes in one micrograph the main message behind this study: as a result of spontaneous vesicle formation and solute encapsulation processes, empty and filled vesicles coexist in the same sample, and whereas it is evident that most of the vesicles contain a low number of ferritin molecules, or are empty, a minority of them (<1%) contains, against expectations, a very high number of ferritin molecules in non-aggregate state. The existence of these “super-filled” vesicles is considered almost impossible according to Poisson statistics, whereas is correctly predicted by the power law. Moreover, in the case of very small vesicles, exceptionally high intravesicle ferritin concentrations have been observed (up to about 300 μM), roughly corresponding to crowding concentrations in biological cells.


Physical Routes to Primitive Cells: An Experimental Model Based on the Spontaneous Entrapment of Enzymes inside Micrometer-Sized Liposomes.

D'Aguanno E, Altamura E, Mavelli F, Fahr A, Stano P, Luisi PL - Life (Basel) (2015)

(a) Ferritin occupancy distribution as determined by analyzing about 7700 individual vesicles imaged by cryo-TEM. Experimental points shown as colored circles, theoretical expectations as empty circles connected by a line; (b) Two conventional POPC vesicles: one is super-filled with ferritin (encapsulating much more ferritin molecules than expected), the other does not contain any ferritin molecule (also against expectations). Images reproduced from [10] with the permission of Wiley.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4390888&req=5

life-05-00969-f004: (a) Ferritin occupancy distribution as determined by analyzing about 7700 individual vesicles imaged by cryo-TEM. Experimental points shown as colored circles, theoretical expectations as empty circles connected by a line; (b) Two conventional POPC vesicles: one is super-filled with ferritin (encapsulating much more ferritin molecules than expected), the other does not contain any ferritin molecule (also against expectations). Images reproduced from [10] with the permission of Wiley.
Mentions: Intrigued by these unexpected conclusions, we then started a direct investigation on solute encapsulation inside spontaneously formed lipid vesicles. We recalled our previous work on the use of the protein ferritin as a marker for the vesicle lumen [4,5]. Individual ferritin molecules contain high amount of iron in form of hydrous ferric oxide phosphate, and it is widely used in electronmicroscopy. Since the number of ferritin molecules inside vesicles can be directly counted in images obtained by cryo-transmission electronmicroscopy (cryo-TEM), and the vesicle size is measurable as well, it is possible to measure the solute occupancy distribution f(n) in vesicle populations and compare it with the expected Poisson distribution, p(n). As shown in Figure 4A, the observed distribution is quite different than the theoretical one. The distribution is not bell-shaped, and—at high n—the measured distribution has a long “tail” of values that are significantly higher than the vanishing small values of the Poisson curve. The experimentally determined distribution strongly resemble a power law, i.e., f(n) ~ 1/na (a > 0), and Figure 4B summarizes in one micrograph the main message behind this study: as a result of spontaneous vesicle formation and solute encapsulation processes, empty and filled vesicles coexist in the same sample, and whereas it is evident that most of the vesicles contain a low number of ferritin molecules, or are empty, a minority of them (<1%) contains, against expectations, a very high number of ferritin molecules in non-aggregate state. The existence of these “super-filled” vesicles is considered almost impossible according to Poisson statistics, whereas is correctly predicted by the power law. Moreover, in the case of very small vesicles, exceptionally high intravesicle ferritin concentrations have been observed (up to about 300 μM), roughly corresponding to crowding concentrations in biological cells.

Bottom Line: We have reported that proteins and ribosomes can be encapsulated very efficiently, against statistical expectations, inside a small number of liposomes.Moreover the transcription-translation mixture, which realistically mimics a sort of minimal metabolic network, can be functionally reconstituted in liposomes owing to a self-concentration mechanism.Here we firstly summarize the recent advancements in this research line, highlighting how these results open a new vista on the phenomena that could have been important for the formation of functional primitive cells.

View Article: PubMed Central - PubMed

Affiliation: Science Department, Roma Tre University, Viale G. Marconi 446, I-00146 Rome, Italy. Alfred.Fahr@uni-jena.de.

ABSTRACT
How did primitive living cells originate? The formation of early cells, which were probably solute-filled vesicles capable of performing a rudimentary metabolism (and possibly self-reproduction), is still one of the big unsolved questions in origin of life. We have recently used lipid vesicles (liposomes) as primitive cell models, aiming at the study of the physical mechanisms for macromolecules encapsulation. We have reported that proteins and ribosomes can be encapsulated very efficiently, against statistical expectations, inside a small number of liposomes. Moreover the transcription-translation mixture, which realistically mimics a sort of minimal metabolic network, can be functionally reconstituted in liposomes owing to a self-concentration mechanism. Here we firstly summarize the recent advancements in this research line, highlighting how these results open a new vista on the phenomena that could have been important for the formation of functional primitive cells. Then, we present new evidences on the non-random entrapment of macromolecules (proteins, dextrans) in phospholipid vesicle, and in particular we show how enzymatic reactions can be accelerated because of the enhancement of their concentration inside liposomes.

No MeSH data available.


Related in: MedlinePlus