Proteomimetic zinc finger domains with modified metal‐binding β‐turns

Abstract

The mimicry of protein tertiary folds by chains artificial in backbone chemical composition leads to proteomimetic analogues with potential utility as bioactive agents and as tools, to shed light on biomacromolecule behavior. Notable successes toward such molecules have been achieved; however, as protein structural diversity is vast, design principles must be continually honed as they are applied to new prototype folding patterns. One specific structure where a gap remains in understanding how to effectively generate modified backbone analogues is the metal-binding β-turn found in zinc finger domains. The literature precedent suggests several factors that may act in concert, including the artificial moiety used to modify the turn, the sequence in which it is applied, and modifications present elsewhere in the domain. Here, we report efforts to gain insights into these issues and leverage these insights to construct a zinc finger mimetic with backbone modifications throughout its constituent secondary structures. We first conduct a systematic comparison of four turn mimetics in a common host sequence, quantifying relative efficacy for use in a metal-binding context. We go on to construct a proteomimetic zinc finger domain in which the helix, strands, and turn are simultaneously modified, resulting in a variant with 23% artificial residues, a tertiary fold indistinguishable from the prototype, and a folded stability comparable to the natural backbone on which the variant is based. Collectively, the results reported provide new insights into the effects of backbone modification on the structure and stability of metal-binding domains and help inform the design of metalloprotein mimetics.

Study on the mechanism of zinc finger proteins ROCK1 and DMD in zinc-mediated cognitive impairment caused by aluminum exposure

“…Now, considering the changes in Zn concentrations after Al exposure, it becomes pertinent to discuss its implications on ROCK1 or other ZNF proteins, with a known Zn binding site or any metal-associated site. The binding of metal to these specialized sites often provides structural integrity to the protein, and any change in the concentration or the nature of these ions can lead to a deviation from the protein's native structure (Rao and Horne, 2020). Such deviations can manifest in numerous ways, from misfolding to loss of function or even gain of toxic function (Jacques et al, 2013).…”

Studying Peptide-Metal Ion Complex Structures by Solution-State NMR

“…Short and longer peptides have been used extensively for modeling and understanding different aspects of zinc finger binding, including metal coordination, folding and actual binding [120][121][122]: Cyclic peptides with linear tails showed conformational and thermodynamic stability relative to linear peptides for modeling zinc fingers, and were able to fold into conformations that reproduced the zinc-ribbon fold of zinc fingers and bound Zn(II) better than their linear counterparts [123]. Peptides were used to study secondary structural elements common in zinc fingers and their role in folding and binding zinc: This enabled designing peptides that were modified by 23% of their native residues and still showed tertiary folds and stability on par with the natural backbone of the original zinc finger protein [124].…”

Effects of altered backbone composition on the folding kinetics and mechanism of an ultrafast-folding protein

“…Most work to this end has focused on determining how changes to backbone composition influence folding thermodynamics. 17–25 In contrast, efforts to probe folding kinetics in these systems are rare. In one pioneering example, a series of artificial β-turn mimetics were introduced in a hairpin loop found in the WW domain of the human protein Pin1.…”

“…As part of an ongoing program seeking insights into fundamental folding behavior of protein tertiary structure mimetics, 21,22,24 we recently explored the effects of altered backbone composition on the folded structure and thermodynamic stability of the B domain of protein A from Staphylococcal bacteria (BdpA), which adopts a compact tertiary fold consisting of three antiparallel α-helices. 29 A combination of experimental biophysical analysis and atomistic simulation yielded insights into the folded states, unfolded states, and equilibrium folding thermodynamics of BpdA and heterogeneous-backbone variants in which artificial monomers of different types were introduced in different regions of the domain.…”