Experimental work
I briefly described the work done in the duration of the project.
Work Package 1: Design and synthesis of short peptide building blocks. The design, synthesis, and analysis of our model peptide derivative are summarised in deliverables, which are included separately. Moreover, we have shown that our design can easily be extended to several different combinations of amino acids.
We have synthesized several different peptides conjugates that can be used to form coacervates,
- FFssFF molecules were used as a model compound
- We exchanged the phenylalanine with other hydrophobic amino acids with increasing and decreasing the hydrophobic ability of the sticker residues.
- The spacers/linkers between two hydrophobic stickers residues were also changes in several molecules to see the effect of polarity of spacers on coacervation.
Work package 2: Coacervation and characterization
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The model compound FFssFF and most of the analogue derivatives are water-soluble compounds. We made the 1 to 15 mg/ml concentration as stock solutions in Milli-Q and increased the pH by using NaOH or any buffer of pH 7.0. This immediately converted the transparent solution to turbid solution as shown in fig 4 and turbidity is responsive towards pH. Under the microscope, we observed the coacervates in turbid solution while there were no coacervates in the transparent solution at pH-6.
Work package 3: Reversibility and Encapsulation of the pigments
In most of the compounds, we focus on the spacer of cystamine, which is a hydrophilic linker. The interesting point of this linker is not only the hydrophilicity to drive the liquid-liquid phase separation but also its redox responsive nature plays a role to make the system more prebiotically relevant. We used the reducing agents DTT and TCEP to reduce the disulfide bond which ultimately dissolve the coacervates and turbidity also gone on reduction.
Work package 4: Creation of functional protocells
Coacervates of diphenylalanine dipeptides (FF-ss-FF) have been found to take up a wide range of prebiotically interesting molecules, like short RNAs, ssDNA, and natural pigments. In this WP-4,
In the report, I investigated the promising results of coacervates to vesicles transformation which deviates us to go in this direction before exploring the replication property of the coacervates. One of the analogue derivatives of FFssFF, where we replaced the phenylalanine with tyrosine and tyrosine is well-known amino acid for enzymatic oxidation. The YFsFY compound form the coacervates in a similar way as FFssFF does form but interestingly by adding the oxidizing agents, the coacervates transform into the semipermeable membrane and we could say the internal core of vesicles/shells is still mobile and similar to the coacervates.
For all oral and poster presentations, cluster meetings, and research papers, we acknowledged the Marie Curie Foundation for the great financial support.
