Effects of As(III) Binding on β-Hairpin Structure

Abstract

While arsenic(III) compounds can exert profound toxicological and pharmacological effects, their modes of action and, in particular, the structural consequences of their binding to cysteinyl side chains in proteins, remain poorly understood. To gain an understanding of how arsenic binding influences beta-structure, pairs of cysteines were introduced into a model monomeric beta-hairpin to yield a family of peptides such that coordination occurs either across the strands or within the same strand of the beta-hairpin. Circular dichroism, NMR, UV-vis spectroscopy, and rapid-reaction studies were used to characterize the binding of monomethylarsonous acid or p-succinylamidephenyl arsenoxide (PSAO) to these peptides. Placement of cysteines at non-hydrogen bond (NHB) positions across the beta-hairpin, such that they occupy the same face of the sheet, was found to enhance the structure as assessed by CD. Cross-strand cysteine residues that project on opposite faces close to the termini of the hairpin can still bind arsenic tightly and show modestly increased beta-sheet content. NMR and modeling studies suggest that arsenic can be accommodated at this locus without disrupting the core interactions stabilizing the turn. However, As(III) binding to nonopposed cysteines, or to cysteines at HB and NHB positions along one strand of the hairpin, caused loss of structure. UV-vis titrations show that all these hairpin peptides bind PSAO stoichiometrically with K(d) values from 13 to 106 nM. Further, binding is moderately rapid, with second-order rate constants for association of 10,000-22,000 M(-1) s(-)1 irrespective of the placement of the cysteines within the hairpin and the consequent extent of structural reorganization required as a result of binding. These studies complement recent work with alpha-helices and further demonstrate that capture of a pair of thiols by As(III) may result in significant changes in local secondary structure in the protein targets of these potent bioactive agents.

The role of protein binding of trivalent arsenicals in arsenic carcinogenesis and toxicity

“…As stated in a recent paper on arsenite binding, protein folding and structure which utilized short peptides, ''It appears that the free energy realized from the acquisition of two new As-S bonds overwhelms the structural biases of those dithiol peptides that present initially unfavorable geometries for arsenic binding" [16]. New crosslinked species can have new biological properties and present to the cell new potential energy surfaces.…”

Theoretical calculations of the relative pKa values of some selected aromatic arsonic acids in water using density functional theory

“…[5][6][7] They also are under consideration from the biological activity point of view yet. [8][9][10] Organo arsenic compounds are very useful starting materials with important application in various fields of chemistry. For example Lloyd et al used the aryl arsonic acids as precursors to As(III) and As(I) species as part of their studies into the chemistry and biological activity of Salvarsan which is the historically-important arsenical drug for treatment of syphilis.…”

Identifying important structural characteristics of arsenic resistance proteins by using designed three-stranded coiled coils

“…Helical scaffolds have been the predominant structural motif used for the incorporation of many types of metal-binding sites, including hemes (16), iron-sulfur clusters (17), dinuclear metal centers (18), copper centers (19,20), and heavy metal-binding trigonal thiolate sites (7,21). ␤-Hairpins and other globular structures that bind As(III), Zn(II), and other metal cofactors have also been designed (22)(23)(24)(25).Despite the increasing number of reports on the design of metallopeptides, detailed structural characterization of many of these systems remains elusive. The class of designed metallopeptides that has been structurally characterized most thor-…”