Amyloid peptides incorporating a core sequence from the amyloid beta peptide and gamma amino acids: relating bioactivity to self-assembly

A series of heptapeptides comprising the core sequence Ab(16–20), KLVFF, of the amyloid b peptide coupled with paired N-terminal c-amino acids are investigated in terms of cytotoxicity reduction and binding to the full Ab peptide, both pointing to inhibition of fibrillisation for selected compounds. This is related to the self-assembly capacity of the heptapeptides.

Three dimensional picture of the local structure of 1,4-Polybutadiene from a complete atomistic model and neutron scattering data

An efficient method of combining neutron diffraction data over an extended Q range with detailed atomistic models is presented. A quantitative and qualitative mapping of the organization of the chain conformation in both glass and liquid phase has been performed. The proposed structural refinement method is based on the exploitation of the intrachain features of the diffraction pattern by the use of internal coordinates for bond lengths, valence angles and torsion rotations. Models are built stochastically by assignment of these internal coordinates from probability distributions with limited variable parameters. Variation of these parameters is used in the construction of models that minimize the differences between the observed and calculated structure factors. A series of neutron scattering data of 1,4-polybutadiene at the region 20320 K is presented. Analysis of the experimental data yield bond lengths for C-C and C=C of 1.54 and 1.35 Å respectively. Valence angles of the backbone were found to be at 112 and 122.8 for the CCC and CC=C respectively. Three torsion angles corresponding to the double bond and the adjacent R and β bonds were found to occupy cis and trans, s(, trans and g( and trans states, respectively. We compare our results with theoretical predictions, computer simulations, RIS models, and previously reported experimental results.

Fibrils and nanotubes assembled from a modified amyloid-β peptide fragment differ in the packing of the same β-sheet building blocks

The peptide AAKLVFF assembles into fibrils in water and nanotubes in methanol. Solid-state NMR data are consistent with fibrils constructed from β-sheet bilayers and nanotubes bounded by a wall of offset β-sheet monolayers. Remarkably distinct morphologies are thus traced to subtle differences in the arrangement of the same fundamental building blocks.

Control of DNA conformation using 3'-S-phosphorothiolate-modified linkages

An in-depth study into the incorporation of multiple 3'-S-phosphorothiolate modifications into oligodeoxynucleotides (ODNs) and their subsequent effect on ODN/DNA and ODN/RNA duplex stability. 3'-S-Phosphorothiolate linkages increase the stability of ODN/RNA duplexes and decrease the stability of ODN/DNA duplexes.

Extended Kalman filtering for robotic proximity sensing

The authors address the problems in using a fiber-optic proximity sensor to detect robot end-effector positioning errors in performing a contact task when uncertainties about target position exist. An extended Kalman filter approach is employed to solve the nonlinear filtering problem. Some experimental results are given.

Synthesis and incorporation into cyclic peptides of tolan amino acids and their hydrogenated congeners: construction of an array of A–B-loop mimetics of the Cε3 domain of human IgE

The disruption of the human immunolobulin E–high affinity receptor I (IgE–FcεRI) protein–protein interaction (PPI) is a validated strategy for the development of anti asthma therapeutics. Here, we describe the synthesis of an array of conformationally constrained cyclic peptides based on an epitope of the A–B loop within the Cε3 domain of IgE. The peptides contain various tolan (i.e., 1,2-biarylethyne) amino acids and their fully and partially hydrogenated congeners as conformational constraints. Modest antagonist activity (IC50 660 μM) is displayed by the peptide containing a 2,2′-tolan, which is the one predicted by molecular modeling to best mimic the conformation of the native A–B loop epitope in IgE.

RACORO extended-term, aircraft observations of boundary-layer clouds

A first-of-a-kind, extended-term cloud aircraft campaign was conducted to obtain an in-situ statistical characterization of continental boundary-layer clouds needed to investigate cloud processes and refine retrieval algorithms. Coordinated by the Atmospheric Radiation Measurement (ARM) Aerial Facility (AAF), the Routine AAF Clouds with Low Optical Water Depths (CLOWD) Optical Radiative Observations (RACORO) field campaign operated over the ARM Southern Great Plains (SGP) site from 22 January to 30 June 2009, collecting 260 h of data during 59 research flights. A comprehensive payload aboard the Center for Interdisciplinary Remotely-Piloted Aircraft Studies (CIRPAS) Twin Otter aircraft measured cloud microphysics, solar and thermal radiation, physical aerosol properties, and atmospheric state parameters. Proximity to the SGP's extensive complement of surface measurements provides ancillary data that supports modeling studies and facilitates evaluation of a variety of surface retrieval algorithms. The five-month duration enabled sampling a range of conditions associated with the seasonal transition from winter to summer. Although about two-thirds of the cloud flights occurred in May and June, boundary-layer cloud fields were sampled under a variety of environmental and aerosol conditions, with about 77% of the flights occurring in cumulus and stratocumulus. Preliminary analyses illustrate use of these data to analyze cloud-aerosol relationships, characterize the horizontal variability of cloud radiative impacts, and evaluate surface-based retrievals. We discuss how an extended-term campaign requires a simplified operating paradigm that is different from that used for typical, short-term, intensive aircraft field programs.

Endosomal proteolysis regulates calcitonin gene-related peptide responses in mesenteric arteries

Background and Purpose: Calcitonin gene‐related peptide (CGRP) is a potent vasodilator, implicated in the pathogenesis of migraine. CGRP activates a receptor complex comprising, calcitonin receptor‐like receptor (CLR) and receptor activity‐modifying protein 1 (RAMP1). In vitro studies indicate recycling of CLR•RAMP1 is regulated by degradation of CGRP in early endosomes by endothelin‐converting enzyme‐1 (ECE‐1). However, it is not known if ECE‐1 regulates the resensitization of CGRP‐induced responses in functional arterial tissue. Experimental Approach: CLR, ECE‐1a‐d and RAMP1 expression in rat mesenteric artery smooth muscle cells (RMA‐SMCs) and mesenteric arteries was analyzed by RT‐PCR and by immunofluorescence and confocal microscopy. CGRP‐induced signaling in cells was examined by measuring cAMP production and ERK activation. CGRP‐induced relaxation of arteries was measured by isometric wire myography. ECE‐1 was inhibited using the specific inhibitor, SM‐19712. Key Results: RMA‐SMCs and arteries contained mRNA for CLR, ECE‐1a‐d and RAMP1. ECE‐1 was present in early endosomes of RMA‐SMCs and in the smooth muscle layer of arteries. CGRP induced endothelium‐independent relaxation of arteries. ECE‐1 inhibition had no effect on initial CGRP‐induced responses but reduced cAMP generation in RMA‐SMCs and vasodilation in mesenteric arteries responses to subsequent CGRP challenges. Conclusions and Implications: ECE‐1 regulates the resensitization of responses to CGRP in RMA‐SMCs and mesenteric arteries. CGRP‐induced relaxation does not involve endothelium‐derived pathways. This is the first report of ECE‐1 regulating CGRP responses in SMCs and arteries. ECE‐1 inhibitors may attenuate an important vasodilatory pathway, implicated in primary headaches and may represent a new therapeutic approach for the treatment of migraine.

Self-assembly of a peptide amphiphile containing l-carnosine and its mixtures with a multilamellar vesicle forming lipid

The self-assembly of the peptide amphiphile (PA) hexadecyl-(β-alaninehistidine) is examined in aqueous solution, along with its mixtures with multilamellar vesicles formed by DPPC (dipalmitoyl phosphatidylcholine). This PA, denoted C16-βAH, contains a dipeptide headgroup corresponding to the bioactive molecule L-carnosine. It is found to selfassemble into nanotapes based on stacked layers of molecules. Bilayers are found to coexist with monolayers in which the PA molecules pack with alternating up−down arrangement so that the headgroups decorate both surfaces. The bilayers become dehydrated as PA concentration increases and the number of layers in the stack decreases to produce ultrathin nanotapes comprised of 2−3 bilayers. Addition of the PA to DPPC multilamellar vesicles leads to a transition to well-defined unilamellar vesicles. The unique ability to modulate the stacking of this PA as a function of concentration, combined with its ability to induce a multilamellar to unilamellar thinning of DPPC vesicles, may be useful in biomaterials applications where the presentation of the peptide function at the surface of self-assembled nanostructures is crucial.

Conductance of amyloid β based peptide filaments: structure–function relations.

Controlling the morphology of self-assembled peptide nanostructures, particularly those based on amyloid peptides, has been the focus of intense research. In order to exploit these structures in electronic applications, further understanding of their electronic behavior is required. In this work, the role of peptide morphology in determining electronic conduction along self-assembled peptide nanofilament networks is demonstrated. The peptides used in this work were based on the sequence AAKLVFF, which is an extension of a core sequence from the amyloid b peptide. We show that the incorporation of a non-natural amino acid, 2-thienylalanine, instead of phenylalanine improves the obtained conductance with respect to that obtained for a similar structure based on the native sequence, which was not the case for the incorporation of 3-thienylalanine. Furthermore, we demonstrate that the morphology of the self-assembled structures, which can be controlled by the solvent used in the assembly process, strongly affects the conductance, with larger conduction obtained for a morphology of long, straight filaments. Our results demonstrate that, similar to natural systems, the assembly and folding of peptides could be of great importance for optimizing their function as components of electronic devices. Hence, sequence design and assembly conditions can be used to control the performance of peptide based structures in such electronic applications.

Collagen or collagen-related peptide cause (Ca2+)i elevation and increased tyrosine phosphorylation in human megakaryocytes.

Since megakaryocytes are the cellular precursors of platelets we have investigated whether they share responses to platelet agonists, in particular collagen. Although previous studies have reported responses to thrombin in non-human megakaryocytes, through studies of single cell calcium responses and protein tyrosine-phosphorylation we demonstrate for the first time that both isolated human megakaryocytes and CD41/61-positive megakaryocytes derived in culture from CD34+ cells share responses to the platelet agonists collagen, collagen-related peptide and thrombin. The responses to either collagen or CRP were seen only in the most mature megakaryocytes and not in megakaryocyte-like cell lines, suggesting that the response to collagen is a characteristic developed late during megakaryocyte differentiation. These primary cells offer the opportunity to use many molecular and cellular techniques to study and manipulate signalling events in response to platelet receptor agonists, which cannot be performed in the small, anucleate platelet itself.

A collagen-like peptide stimulates tyrosine phosphorylation of syk and phospholipase C gamma2 in platelets independent of the integrin alpha2beta1.

Activation of platelets by collagen is mediated through a tyrosine kinase-dependent pathway that is associated with phosphorylation of the Fc receptor gamma chain, the tyrosine kinase syk, and phospholipase C gamma2 (PLC gamma2). We recently described a collagen-related triple-helical peptide (CRP) with the sequence GCP*(GPP*)GCP*G (single letter amino acid code: P* = hydroxyproline; Morton et al, Biochem J306:337, 1995). The cross-linked peptide is a potent stimulus of platelet activation but, unlike collagen, does not support alpha2beta1-mediated, Mg2+-dependent adhesion, suggesting that its action is independent of the integrin alpha2beta1. This finding suggests the existence of a platelet receptor other than alpha2beta1 that underlies activation. In the present study, we show that CRP stimulates tyrosine phosphorylation of the same pattern of proteins in platelets as collagen, including syk and PLC gamma2. Protein tyrosine phosphorylation induced by CRP is not altered in the absence of Mg2+ or the presence of monoclonal antibodies (MoAbs) to the integrin alpha2beta1 (MoAb 6F1 and MoAb 13), conditions that prevent the interaction of collagen with the integrin. In contrast, phosphorylation of syk and PLC gamma2 by collagen is partially reduced by MoAb 6F1 and MoAb 13 or by removal of Mg2+. This may reflect a direct role of alpha2beta1 in collagen-induced signaling events or an indirect role in which the integrin facilitates the binding of collagen to its signaling receptor. The results show an alpha2beta1-independent pathway of platelet activation by CRP that involves phosphorylation of syk and PLC gamma2. This pathway appears to contribute to platelet activation by collagen.

Altering peptide fibrillization by polymer conjugation

A strategy is presented that exploits the ability of synthetic polymers of different nature to disturb the strong selfassembly capabilities of amyloid based β-sheet forming peptides. Following a convergent approach, the peptides of interest were synthesized via solid-phase peptide synthesis (SPPS) and the polymers via reversible addition−fragmentation chain transfer (RAFT) polymerization, followed by a copper(I) catalyzed azide− alkyne cycloaddition (CuAAC) to generate the desired peptide− polymer conjugates. This study focuses on a modified version of the core sequence of the β-amyloid peptide (Aβ), Aβ(16−20) (KLVFF). The influence of attaching short poly(Nisopropylacrylamide) and poly(hydroxyethylacrylate) to the peptide sequences on the self-assembly properties of the hybrid materials were studied via infrared spectroscopy, TEM, circular dichroism and SAXS. The findings indicate that attaching these polymers disturbs the strong self-assembly properties of the biomolecules to a certain degree and permits to influence the aggregation of the peptides based on their β-sheets forming abilities. This study presents an innovative route toward targeted and controlled assembly of amyloid-like fibers to drive the formation of polymeric nanomaterials.

Reversible thermal transition of polydiacetylene based on KTTKS collagen sequence

Here we explore the physico-chemical properties of a peptide amphiphile obtained by chemical conjugation of the collagenstimulating peptide KTTKS with 10,12-pentacosadiynoic acid which photopolymerizes as a stable and extended polydiacetylene. We investigate the self-assembly of this new polymer and rationalize its peculiar behavior in terms of a thermal conformational transition. Surprisingly, this polymer shows a thermal transition associated with a non-cooperative increase in b-sheet content at high temperature.