Source-based synthetic examples
Keep the near neighbor that can challenge the mechanism.
In a retained synthetic sequence example, the target and paralog A carry a cysteine at the aligned site; paralog B carries serine. The presence check identifies A as the immediate counterscreen priority. A separate mutation model then asks whether local chemistry supports a selectivity story: adding a negative charge 6 Å from the cysteine raises its modeled pKa and lowers its reactive thiolate fraction under the stated assumptions.
- Cys / Cys / SerCompare the aligned site
Target and paralog A share the handle. Paralog B substitutes serine.
- −1 charge, 6 ÅChallenge the local chemistry
In the separate toy electrostatic model, pKa rises and thiolate availability falls.
- Prioritize AChoose the matched counterscreen
Test engagement against the conserved paralog; test mutant and original protein separately.
- What the sequence result gives you · One of two supplied paralogs retains the cysteine. That is a concrete reason to test that paralog, while keeping broader off-target and recognition questions open.
- What the mutation check adds · The toy model uses pH 7.4, an initial pKa of 8.9 and effective dielectric 20. Its predicted direction challenges an assumed reactivity advantage. Measure recognition and reaction separately in the matched proteins before treating the shift as useful selectivity.
From sequence difference to reaction hypothesis
A sequence alignment identifies conserved and substituted residues. The BioTwin assessment adds modeled structural and electrostatic context around a proposed handle. A residue can be present in two proteins while its environment differs; absence alone also leaves other off-target routes to consider.
Make the comparison experimental
Current output is a computational selectivity hypothesis. Choose relevant paralogs and variants for a matched counterscreen, separating recognition, reaction rate and exposure when they affect interpretation. Neither a non-conserved handle nor a modeled thiolate change proves selective engagement.
Selectivity has more than one source
Local electrostatic design could change target chemistry, while plasma competition could defeat it before engagement. Combining those questions may reveal why a favorable target/paralog comparison fails in a broader setting. Each component and its inputs need to be checked.
- Local electrostatics · Compare recognition, reaction conditions and exposure to prioritize candidate designs and their next laboratory tests. Local electrostatics
- Competing reactions · Follow the extracellular loss processes that may dominate capture. Competing reactions