title: Evolutionary bridges to new protein folds: design of C-terminal Cro protein chameleon sequences.
abstract: 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 λ(WDD)-SASF2 show moderate thermal stability in denaturation curves monitored by circular dichroism (T(m) values of 46 and 55°C, respectively), while P22-SASF3 and λ(WDD)-SASF3 have somewhat reduced stability (T(m) values of 33 and 49°C, respectively). (13)C and (1)H 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 λ(WDD)-SASF2 (residues 40-45 and 50-52), 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 (15)N-(1)H 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.
authors.affiliation: Department of Chemistry and Biochemistry, University of Arizona, Tucson, AZ 85721-0088, USA.
authors.forename: William J
authors.initials: WJ
authors.lastname: Anderson
authors.forename: Laura O
authors.initials: LO
authors.lastname: Van Dorn
authors.forename: Wendy M
authors.initials: WM
authors.lastname: Ingram
authors.forename: Matthew H J
authors.initials: MH
authors.lastname: Cordes
mesh_terms: D000595:Amino Acid Sequence; D019143:Evolution, Molecular; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D017510:Protein Folding; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D054334:Viral Regulatory and Accessory Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D012097:Repressor Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins