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ABSTRACT This study focuses on investigating the conformational structure and zinc(II) affinity of a zinc finger‐like motif (ZFM) peptide with the sequence acetyl‐His1‐Cys2‐Gly3‐Pro4‐Gly5‐His6‐Cys7, where bold highlights the potential zinc(II) binding sites. Zinc fingers are crucial protein motifs known for their high specificity and affinity for zinc ions. The ZFM peptide's sequence contains the 2His‐2Cys zinc‐binding sites similar to those in natural zinc finger proteins but without the hydrophobic core, making it a valuable model for studying zinc(II)–peptide interactions. Previous research on related peptides showed that collision cross sections and B3LYP modeling predicted that the His‐2Cys‐carboxyl terminus coordination of zinc(II) was more stable than the 2His‐2Cys. Employing a comprehensive approach integrating ion mobility–mass spectrometry and theoretical modeling techniques, various zinc(II) binding modes of the ZFM have been thoroughly compared to ascertain their influence on the competitive threshold collision‐induced dissociation method for measuring the relative gas‐phase Zn(II) affinity of the ZFM peptide. The measured Zn(II) affinity of ZFM is greater than those measured recently for two peptides with similar primary structures, acetyl‐His1‐Cys2‐Gly3‐Pro4‐Gly5‐Gly6‐Cys7and acetyl‐Asp1‐His2‐Gly3‐Pro4‐Gly5‐Gly6‐Cys7, indicating the preference for the His1‐Cys2‐His6‐Cys7side groups for coordinating zinc(II) over the His‐2Cys‐carboxyl terminus or Asp‐His‐Cys‐carboxyl terminus in these related heptapeptides.
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This content will become publicly available on July 1, 2026
Effect of the Tyr5 and His6 substituent groups on the zinc affinities and conformational structures of the acetyl-His1-Cys2-Gly3-Pro4-X5-X6-Cys7 heptapeptides
In this study, we investigate the influence of the Tyr5 and His6 substituent groups on the zinc-binding affinities and conformational properties of a series of acetylated heptapeptides, acetyl-His1-Cys2-Gly3-Pro4-X5-X6-Cys7 focusing on the impacts where X5-X6 are either Tyr5-Gly6, Tyr5-His6, Gly5-Gly6, or Gly5-His6. Utilizing traveling-wave ion mobility-mass spectrometry and molecular modeling techniques we analyze the zinc binding interactions and peptide coordination behavior. The zinc binding peptides (ZBPs) relative zinc affinities were measured across pH 5 to pH 10 by monitoring the solution-phase formation of the [ZBP+Zn(II)] complex by utilizing native MS in negative ion mode to preserve the solution-phase binding of Zn(II) to the peptides. Furthermore, their relative gas-phase Zn(II) affinities were measured using competitive threshold collision-induced dissociation (TCID) of the [ZBP+Zn(II)+NTA] complex, by modeling the two competing dissociation channels: [ZBP+Zn(II)]- + NTA or [Zn(II)+NTA] + ZBP, where NTA is nitrilotriacetic acid. Our examinations also tested whether there was an effect of the formation of the [ZBP+Zn(II)+NTA] complexes from solutions at different pHs, before they are electrosprayed into the gas-phase for the TCID analyses. Both solution- and gas-phase measurements predicted the heptapeptide with the Gly5-His6 residues had the greatest zinc affinity and that the presence of Tyr5 and His6 altered the zinc affinity and induced distinct conformational changes due to changes in the coordination of the zinc. This research enhances our understanding of zinc-peptide interactions, with implications for the design of peptide-based metalloproteins, which may guide the design of novel ZBPs for therapeutic, biotechnological or environmental remediation applications.
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- Award ID(s):
- 2247511
- PAR ID:
- 10604014
- Publisher / Repository:
- Elsevier
- Date Published:
- Journal Name:
- International Journal of Mass Spectrometry
- Volume:
- 513
- Issue:
- C
- ISSN:
- 1387-3806
- Page Range / eLocation ID:
- 117458
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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