Ca²⁺‐induced conformational transitions of phosphorylated peptides

Abstract

CD spectroscopic studies on protected peptides containing lysine and serine, or phosphoserine, and on serine-containing fragments of the neurofilament protein midsized subunit, both in the unphosphorylated and phosphorylated form, are reported. The introduction of the phosphoryl group was not found to have a significant spectral effect in aqueous solution. In trifluoroethanol (TFE), spectral shifts toward unordered (type U) spectra or the appearance of distorted spectra likely reflect the adoption of aperiodic polypeptide conformations due to salt bridge(s) between negatively charged phosphoserine and positive lysine side-chain groups. A turn-stabilizing effect of phosphorylation was also observed. CD-monitored titration experiments in TFE revealed a high conformational sensitivity of phosphopeptides toward Ca2+ ions. The appearance of the unordered spectra or spectral shifts were the sign of a bulk disordering effect of Ca2+ ions. Spectra with specific spectroscopic features reflect the formation of Ca2+ complexes and the adoption of ordered unique backbone conformations. When ordered structures were obtained on addition of Ca2+ ions, the observed CD curves showed a resemblance to the spectrum of beta-pleated sheets. This may originate from chain extension and the formation of beta-pleated sheet segments fixed by Ca2+ bridges between PO3H-1 groups of adjacent peptide chains. The data clearly show that the effect of the Ca2+ ions is highly specific: the sequence, chain length, presence and distribution of charged side-chain groups, degree and site of phosphorylation, and environmental factors appear to be determining in the process of chain extension or beta-sheet formation.

A Conformationally Stable Acyclic β‐Hairpin Scaffold Tolerating the Incorporation of Poorly β‐Sheet‐Prone Amino Acids

“…The resulting peptide 1 is not well folded, despite its still high similarity with the Trp K pocket peptide mentioned above: in detail, the CD spectrum of the latter [16] is characterized by an intense minimum close to 215 nm, indicative of β‐sheet structure, as well as by a moderate maximum close to 230 nm attributed to exciton interactions between the aromatic side chains [18] . Instead, the CD spectrum of peptide 1 features a negative contribution at 215 nm, accompanied by a broad minimum below 200 nm, which is characteristic of unordered or irregular conformations [19] (Figure 2a). The addition of secondary structure‐stabilizing cosolvents, [20] like 2,2,2‐trifluoroethanol (TFE) and MeOH, did not help peptide 1 in adopting the fold of the Trp K pocket peptide: in fact, MeOH barely affected the shape of the CD curve, whereas TFE led to a minimum at 205 nm with a negative shoulder close to 215 nm and a maximum close to 190 nm, probably reflecting the formation of type I and III β‐turns [21]…”

The solubilization of model Alzheimer tangles: reversing the beta-sheet conformation induced by aluminum with silicates.

“…Using circular dichroism (CD) to monitor conformational changes, the 13-and 17-amino acid peptides [NF-M13, GluGlu-Lys-Gly-Lys-Ser-Pro-Val-Pro-Lys-Ser-Pro-Val-GluGlu-Lys-Gly (NF-M17), [EEKG]NF-M13] have been titrated with metal ions. Both Ca2+ and A13+ induce 13-sheet formation before causing the peptide to precipitate (19,20); this is the identical conformation found in plaques (21). Attempts to reverse the effect of the bound metal have been reported.…”

Stable intrachain and interchain complexes of neurofilament peptides: a putative link between Al3+ and Alzheimer disease.

“…2A). The importance of electrostatic interactions is emphasized by the observation that NF-M17 shows characteristic (3-sheet shifts of the CD spectra in TFE upon addition of increasing amounts of Ca2+, whereas NF-M17-NH2, lacking the C-terminal negatively charged carboxylate, does not (15,16).…”

“…The In an attempt to find a molecular mechanism for the effect of aluminum to induce aggregation of neuronal proteins or their catabolic fragments, circular dichroism (CD) spectroscopic studies have been performed in our laboratories on synthetic fragments of human neurofilament mid-sized subunit protein (NF-M), KSPVPKSPVEEKG (NF-M13), and EEKGKSPVPKSPVEEKG (NF-M17), as well as on their alanine-substituted and/or serine-phosphorylated derivatives (15)(16)(17). CD spectra were recorded as described (18) and deconvoluted into secondary structural components by the convex constraint analysis method (19,20 and A13+ were found to induce the adoption of the (3-pleated sheet conformation in the C terminus in a low-dielectricconstant environment, in 2,2,2-trifluoroethanol (TEE).…”

“…The 13-mer peptide NF-M13 is repeated six times at the C terminus of NF-M, and the KSPV tetrapeptide sequence is a potential phosphorylation site of both NF-M and tau protein (21). On the basis of CD (15,16) Fourier-transform infrared spectroscopic (unpublished data), and chemical (15) evidence, Ca2+ and A13+ were found to induce the adoption of the (3-pleated sheet conformation in the C terminus in a low-dielectricconstant environment, in 2,2,2-trifluoroethanol (TEE). These effects were absent in aqueous medium.…”