5 ms·

I would have imagined labs these days just synthesise the mRNA and then let a ribosome do it’s work?

dnautics 5y ago
You can't always do that. Suppose you want a peptide with an unnatural amino acid... Or a natural amino acid with a specific post translational modification...
- XorNot 5y ago
  Unnatural amino acids can be done though - there's been a bunch of work on getting/finding ribosomes which will polymerize all sorts of whacky stuff.
  - dnautics 5y ago
    I'm aware, I used to work next to Floyd Romesberg. Yields are low, suppose you want to put TWO strange things in. Suppose you want to build a depsipeptide (or a dipeptide analog with C=C linkage). Suppose you need a 12-mer (ribosomes are very bad at short peptides).
    - cing 5y ago
      Not saying it's easy but ribosomal synthesis of consecutive non-canonical amino acids has been achieved by some groups (https://www.cell.com/cell-chemical-biology/fulltext/S2451-9456(16)30435-4 https://www.cell.com/cell-chemical-biology/fulltext/S2451-94..., https://pubs.acs.org/doi/10.1021/jacs.8b07247 https://pubs.acs.org/doi/10.1021/jacs.8b07247), and many of these peptides are extremely short. Figure 5 of this manuscript describes successful applications of this for hit identification often in the context of massive peptide library screens, https://pubs.acs.org/doi/10.1021/acs.accounts.1c00391 https://pubs.acs.org/doi/10.1021/acs.accounts.1c00391
      - dnautics 5y ago
        it's not really a generalizable solution, though, because suppose you need an unnatural AA that needs a specialized ribosomal cavity and that cavity hasn't been developed yet. And you need the material by a year's time. You are not going to evolve a ribosome to make your peptide. And, there are post-translational modifications that are too big to fit into ANY cavity, though in theory you could use tricksy chemistry to figure out a downstream processing step, that chemistry may not exist yet, or it may require a pure enzyme that is a pain in the ass to obtain, etc. etc. Finally, charging the correct tRNA is not necessarily easy or efficient. Unlike well-designed programming architectures, biology is usually not "trivially composable". Oddly, SPPS is. Yes, there are corner cases, but those are more the exception than the rule.
asdff 5y ago
even better is to put it in yeast or e coli and let them be your factory

You can't always do that. Suppose you want a peptide with an unnatural amino acid... Or a natural amino acid with a specific post translational modification...

XorNot 5y ago
Unnatural amino acids can be done though - there's been a bunch of work on getting/finding ribosomes which will polymerize all sorts of whacky stuff.
- dnautics 5y ago
  I'm aware, I used to work next to Floyd Romesberg. Yields are low, suppose you want to put TWO strange things in. Suppose you want to build a depsipeptide (or a dipeptide analog with C=C linkage). Suppose you need a 12-mer (ribosomes are very bad at short peptides).
  - cing 5y ago
    Not saying it's easy but ribosomal synthesis of consecutive non-canonical amino acids has been achieved by some groups (https://www.cell.com/cell-chemical-biology/fulltext/S2451-9456(16)30435-4 https://www.cell.com/cell-chemical-biology/fulltext/S2451-94..., https://pubs.acs.org/doi/10.1021/jacs.8b07247 https://pubs.acs.org/doi/10.1021/jacs.8b07247), and many of these peptides are extremely short. Figure 5 of this manuscript describes successful applications of this for hit identification often in the context of massive peptide library screens, https://pubs.acs.org/doi/10.1021/acs.accounts.1c00391 https://pubs.acs.org/doi/10.1021/acs.accounts.1c00391
    - dnautics 5y ago
      it's not really a generalizable solution, though, because suppose you need an unnatural AA that needs a specialized ribosomal cavity and that cavity hasn't been developed yet. And you need the material by a year's time. You are not going to evolve a ribosome to make your peptide. And, there are post-translational modifications that are too big to fit into ANY cavity, though in theory you could use tricksy chemistry to figure out a downstream processing step, that chemistry may not exist yet, or it may require a pure enzyme that is a pain in the ass to obtain, etc. etc. Finally, charging the correct tRNA is not necessarily easy or efficient. Unlike well-designed programming architectures, biology is usually not "trivially composable". Oddly, SPPS is. Yes, there are corner cases, but those are more the exception than the rule.

asdff 5y ago

even better is to put it in yeast or e coli and let them be your factory