* Two foldons make double-CI2 become non-two-state refolding. Stable intermediate accumulated in the refolding process.

* The result showed that LA49(I)+WT(II) and AG16(I)+WT(II) were described by two-state transition, and they did not populate a stable intermediate.

* The conformers NA and NB have different unfolding and refolding rate. And thes two reactions are likely to proceed in parallel, not sequentially.

* This work is of interest from the viewpoints of protein engineering and molecular evolution.

* Side-chain conformers make protein engineering still challenged.
Comformers ---> evolution premature!