A time-dependent fluorescence shift method, biomimetic colorimetric assays, and molecular dynamics simulations have been performed in search of explanations why arginine rich peptides with intermediate lengths of about 10 amino acids translocate well through cellular membranes, while analogous lysine rich peptides do not. First, we demonstrate that an important factor for efficient peptide adsorption, as the first prerequisite for translocation across the membrane, is the presence of negatively charged phospholipids in the bilayer. Second, we observe a strong tendency of adsorbed arginine (but not lysine) containing peptides to aggregate at the bilayer surface. We suggest that this aggregation of oligoarginines leads to partial disruption of the bilayer integrity due to the accumulated large positive charge at its surface, which increases membrane-surface interactions due to the increased effective charge of the aggregates. As a result, membrane penetration and translocation of medium length oligoarginines becomes facilitated in comparison to single arginine and very long polyarginines, as well as to lysine containing peptides.

The development of selective agents capable of discriminating between protein kinase C (PKC) isoforms and other diacylglycerol (DAG)-responsive C1 domain-containing proteins represents an important challenge. Recent studies have highlighted the role that Ras guanine nucleotide-releasing protein (RasGRP) isoforms play both in immune responses as well as in the development of prostate cancer and melanoma, suggesting that the discovery of selective ligands could have potential therapeutic value. Thus far, the N-methyl-substituted indololactone 1 is the agonist with the highest reported potency and selectivity for RasGRP relative to PKC. Here we present the synthesis, binding studies, cellular assays and biophysical analysis of interactions with model membranes of a family of regioisomers of 1 (compounds 2 to 5) that differ in the position of the linkage between the indole ring and the lactone moiety. These structural variations were studied to explore the interaction of the active complex (C1 domain-ligand) with cellular membranes, which is believed to be an important factor for selectivity in the activation of DAG-responsive C1 domain containing signaling proteins. All compounds were potent and selective activators of RasGRP when compared to PKCα with selectivities ranging from 6 to 65 fold. However, the parent compound 1 was appreciably more selective than any of the other isomers. In intact cells, modest differences in the patterns of translocation of the C1 domain targets were observed. Biophysical studies using giant vesicles as model membranes did show substantial differences in terms of molecular interactions impacting lipid organization, dynamics and membrane insertion. However, these differences did not yield correspondingly large changes in patterns of biological response, at least for the parameters examined.

This paper introduces a simple color assay for detection of hypercholesterolemia and monitoring the effect of therapies directed at mitigating this physiological condition. The colorimetric system might constitute a novel platform for assessing patient vulnerability towards the development of atherosclerosis.

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Evaluation of the extent of lipoprotein oxidation and the effects of therapeutic and dietary agents upon lipid peroxidation processes are critical in understanding and combating atherosclerotic diseases. Bioanalytical chromatic assays based upon polydiacetylene (PDA) have recently emerged as a useful platform for analysis of lipoprotein oxidation. This article describes the principles of PDA-based sensing, and its application to the study of lipids peroxidation, both in purified lipoprotein samples, as well as in blood plasma specimens.