Recently, much attention has been given to the problem of drug delivery through the cell-membrane in order to treat and manage several diseases. The discovery of cell penetrating peptides (CPPs) represents a major breakthrough for the transport of large-cargo molecules that may be useful in clinical applications. CPPs are rich in basic amino acids such as arginine and lysine and are able to translocate over membranes and gain access to the cell interior. They can deliver large-cargo molecules, such as oligonucleotides, into cells. Endocytosis and direct penetration have been suggested as the two major uptake mechanisms, a subject still under debate. Unresolved questions include the detailed molecular uptake mechanism(s), reasons for cell toxicity, and the delivery efficiency of CPPs for different cargoes. Here, we give a review focused on uptake mechanisms used by CPPs for membrane translocation and certain experimental factors that affect the mechanism(s).

Metal binding to the amyloid β‐peptide is suggested to be involved in the pathogenesis of Alzheimer's disease. We used high‐resolution NMR to study zinc binding to amyloid β‐peptide 1–40 at physiologic pH. Metal binding induces a structural change in the peptide, which is in chemical exchange on an intermediate rate, between the apo‐form and the holo‐form, with respect to the NMR timescale. This causes loss of NMR signals in the resonances affected by the binding. Heteronuclear correlation experiments, ¹⁵N‐relaxation and amide proton exchange experiments on amyloid β‐peptide 1–40 revealed that zinc binding involves the three histidines (residues 6, 13 and 14) and the N‐terminus, similar to a previously proposed copper‐binding site [Syme CD, Nadal RC, Rigby SE, Viles JH (2004) J Biol Chem279, 18169–18177]. Fluorescence experiments show that zinc shares a common binding site with copper and that the metals have similar affinities for amyloid β‐peptide. The dissociation constant K_d of zinc for the fragment amyloid β‐peptide 1–28 was measured by fluorescence, using competitive binding studies, and that for amyloid β‐peptide 1–40 was measured by NMR. Both methods gave K_d values in the micromolar range at pH 7.2 and 286 K. Zinc also has a second, weaker binding site involving residues between 23 and 28. At high metal ion concentrations, the metal‐induced aggregation should mainly have an electrostatic origin from decreased repulsion between peptides. At low metal ion concentrations, on the other hand, the metal‐induced structure of the peptide counteracts aggregation.

One of the grand challenges of biophysical chemistry is to understand the principles that govern protein misfolding and aggregation, which is a highly complex process that is sensitive to initial conditions, operates on a huge range of length- and timescales, and has products that range from protein dimers to macroscopic amyloid fibrils. Aberrant aggregation is associated with more than 25 diseases, which include Alzheimer's, Parkinson's, Huntington's, and type II diabetes. Amyloid aggregation has been extensively studied in the test tube, therefore under conditions that are far from physiological relevance. Hence, there is dire need to extend these investigations to in vivo conditions where amyloid formation is affected by a myriad of biochemical interactions. As a hallmark of neurodegenerative diseases, these interactions need to be understood in detail to develop novel therapeutic interventions, as millions of people globally suffer from neurodegenerative disorders and type II diabetes. The aim of this review is to document the progress in the research on amyloid formation from a physicochemical perspective with a special focus on the physiological factors influencing the aggregation of the amyloid-β peptide, the islet amyloid polypeptide, α-synuclein, and the hungingtin protein.