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.

A polymetamorphic protein

“…5,18,51 Sequences that can encode two stable folds have been referred to as 'bridges', 'switches', 'metamorphic proteins' and 'transformer proteins' among other terms. 2,15,52-54 S-VLV is in effect a three-way bridge or three-fold switch, 53 capable in principle of acting as a hub for mutational traffic between at least three stable folding patterns related by nontrivial changes in secondary structure.…”

“…14 Limited mutagenesis of small natural proteins populates alternate folds in certain cases, [15][16][17] and accumulation of simple mutations can lead to evolution of new folds. [17][18][19] Some modeling studies have found numerous close approaches or paths between different structures in sequence space [20][21][22][23] and even sequence ''supernetworks'' 24,25 that span many protein folds. In addition, sequences with two folds or functions may confer a fitness advantage under certain models of adaptive evolution.…”

“…28,[33][34][35] High or increased hydrophobicity is experimentally associated with chameleon sequences, lowered structural specificity, or outright fold changes in a number of proteins. 15,18,[36][37][38] Effects can range from increased dynamics and loss of tertiary specificity without a clear change in fold, 36,37 to native state aggregation, 39 to variations in tertiary folding topology, 15 to nonnative aggregation including amyloidogenesis. [40][41][42] In previous work (Fig.…”

Simplification of complexity in protein molecular systems by grouping amino acids: a view from physics

“…Yet, the possible number of proteins is still astronomical. For some sequences, their native structures are conservative with respect to the mutations of amino acids, while some behave like chameleons and may adopt various native conformations under different conditions [36][37][38]. The whole space built with these sequences is not uniform in their foldability and dynamic properties.…”

Sequence-to-Structure Relation in Proteins-Amyloidogenic Proteins with Chameleon Sequences

“…Fragments which exhibit this property are referred to as chameleon sequences [26,27], and their existence is linked to the puzzling phenomenon of "intrinsically disordered" structures [28]. Several databases of chameleon sequences have been published [29,30] and they present an interesting study subject, given the involvement of chameleon sequences in prions and amyloids [31].…”