Evolutionary bridges to new protein folds: Design of C-terminal Cro protein chameleon sequences

William J. Anderson, Laura O. Van Dorn, Wendy M. Ingram, Matthew H.J. Cordes

Research output: Contribution to journalArticlepeer-review

12 Scopus citations


Regions of amino-acid sequence that are compatible with multiple folds may facilitate evolutionary transitions in protein structure. In a previous study, we described a heuristically designed chameleon sequence (SASF1, structurally ambivalent sequence fragment 1) that could adopt either of two naturally occurring conformations (α-helical or β-sheet) when incorporated as part of the C-terminal dimerization subdomain of two structurally divergent transcription factors, P22 Cro and λ Cro. Here we describe longer chameleon designs (SASF2 and SASF3) that in the case of SASF3 correspond to the full C-terminal half of the ordered region of a P22 Cro/λ Cro sequence alignment (residues 34-57). P22-SASF2 and λWDD-SASF2 show moderate thermal stability in denaturation curves monitored by circular dichroism (Tm values of 46 and 55°C, respectively), while P22-SASF3 and λWDD-SASF3 have somewhat reduced stability (Tm values of 33 and 49°C, respectively). 13C and 1H NMR secondary chemical shift analysis confirms two C-terminal α-helices for P22-SASF2 (residues 36-45 and 54-57) and two C-terminal β-strands for λWDD-SASF2 (residues 4045 and 5052), corresponding to secondary structure locations in the two parent sequences. Backbone relaxation data show that both chameleon sequences have a relatively well-ordered structure. Comparisons of 15N- 1H correlation spectra for SASF2 and SASF3-containing proteins strongly suggest that SASF3 retains the chameleonism of SASF2. Both Cro C-terminal conformations can be encoded in a single sequence, showing the plausibility of linking different Cro folds by smooth evolutionary transitions. The N-terminal subdomain, though largely conserved in structure, also exerts an important contextual influence on the structure of the C-terminal region.

Original languageEnglish (US)
Pages (from-to)765-771
Number of pages7
JournalProtein Engineering, Design and Selection
Issue number9
StatePublished - Sep 2011


  • NMR spectroscopy
  • chameleon sequence
  • protein design

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biochemistry
  • Molecular Biology


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