Consecutive Aromatic Residues Are Required for Improved Efficacy of ß-Sheet Breakers.

Alzheimer's disease is a fatal neurodegenerative malady which up to very recently did not have approved therapy modifying its course. After controversial approval of aducanumab (monoclonal antibody clearing ß-amyloid plaques) by FDA for use in very early stages of disease, possibly new avenue opened for the treatment of patients. In line with this approach is search for compounds blocking aggregation into amyloid oligomers subsequently forming fibrils or compounds helping in getting rid of plaques formed by ß-amyloid fibrils. Here we present in silico work on 627 sixtapeptide ß-sheet breakers (BSBs) containing consecutive three aromatic residues. Three of these BSBs caused dissociation of one or two ß-amyloid chains from U-shaped ß-amyloid protofibril model 2BEG after docking and subsequent molecular dynamics simulations. Thorough analysis of our results let us postulate that the first steps of binding these successful BSBs involve π-π interactions with stacked chains of F19 and later also with F20 (F3 and F4 in 2BEG model of protofibril). The consecutive location of aromatic residues in BSBs makes them more attractive for chains of stacked F3 and F4 within the 2BEG model. Spotted by us, BSBs may be prospective lead compounds for an anti-Alzheimer's therapy.

Mechanisms of the modulation of actin-myosin interactions by A1-type myosin light chains.

BACKGROUND: The interaction of N-terminal extension of the myosin A1 essential light chain (A1 ELC) with actin is receiving increasing attention as a target in utilizing synthetic A1 ELC N-terminal-derived peptides in cardiac dysfunction therapy. METHODS: To elucidate the mechanism by which these peptides regulate actin-myosin interaction, here we have investigated their effects on the myosin subfragment 1 (S1)-induced polymerization of G-actin. RESULTS: The MLCFpep and MLCSpep peptides spanning the 3-12 of A1 ELC sequences from fast and slow skeletal muscle, respectively, increased the rate of actin polymerization not only by S1(A2) but also the rate of S1(A1)-induced actin polymerization, suggesting that they did not interfere with the direct binding of A1 ELC with actin. The efficiency of actin polymerization in the presence of the N-terminal ELC peptides depended on their sequence. Substitution of aspartic acid for neutral asparagine at position 5 of MLCFpep dramatically enhanced its ability to stimulate S1-induced polymerization and enabled it to initiate polymerization of G-actin in the absence of S1. CONCLUSIONS: These and other results presented in this work suggest that the modulation of myosin motor activity by N-terminal ELC peptides is exerted through a change in actin filament conformation rather than through blocking the A1 ELC-actin interaction. GENERAL SIGNIFICANCE: The results imply the possibility of enhancing therapeutic effects of these peptides by modifications of their sequence.

ß-sheet breakers with consecutive phenylalanines: Insights into mechanism of dissolution of ß-amyloid fibrils.

ß-sheet breakers (BSB) constitute a class of peptide inhibitors of amyloidogenesis, a process which is a hallmark of many diseases called amyloidoses, including Alzheimer's disease (AD); however, the molecular details of their action are still not fully understood. Here we describe the results of the computational investigation of the three BSBs, iaß6 (LPFFFD), iaß5 (LPFFD), and iaß6_Gly (LPFGFD), in complex with the fibril model of Aß42 and propose the kinetically probable mechanism of their action. The mechanism involves the binding of BSB to the central hydrophobic core (CHC) region (LVFFA) of Aß fibril and the π-stacking of its Phe rings both internally and with the Aß fibril. In the process, the Aß fibril undergoes distortion accumulating on the side of chain A (located on the odd tip of the fibril). In a single replica of extended molecular dynamics run of one of the iaß6 poses, the distortion concludes in a dissociation of chain A from the fibril model of Aß42. Altogether, we postulate that including consecutive Phe residues into BSBs docked around Phe 20 in the CHC region of Aß42 improve their potency for dissolution of fibrils.

Diet as a risk factor for Alzheimer's disease / Dieta jako czynnik ryzyka choroby Alzheimera.

Chronic exposure to an unhealthy diet is one of the causes of civilization diseases and significantly affects the average longevity. The impact of diet on health is extremely complicated due to the chemical diversity of its composition. The diet provides over 26,000 biochemicals and even more of their metabolites. Among this diversity, three macronutrients: proteins, carbohydrates and fats can be identified that provide energy, and in addition providing their metabolites. According to the latest concepts of the impact of macronutrients in diet on human health, their mutual proportions and not solely absolute quantities are of great importance. In our article we present a short discussion of our own research on this problem in relation to the incidence of Alzheimer's disease against the background of contemporary biochemical and epidemiological literature.

Proportions of macronutrients, including specific dietary fats, in prospective anti-Alzheimer's diet.

Here we present a novel life-long whole-population study, which enabled us to predict a diet that, in terms of macronutrient proportions, may be prophylactic against Alzheimer's Disease (AD). The method is based on the existence of oscillations in the correlation between historical per capita personal income (PCPI) and age-adjusted death rates (AADR) for AD for each state of the USA in 2005. These oscillations can be explained by changing proportions of macronutrients in the average American diet between 1929 and 2005. We assumed that reducing future correlation of PCPI with AADR will reduce the population's susceptibility to AD. Based on the results of fitting macronutrient availabilities to the variability of Roriginal, using Generalized Additive Models (GAM) analysis, we constructed four "Calculator" equations. The Calculator allowed for prediction of an optimal diet characterized by low correlation of PCPI with AADR (Rpredicted) and minimum energy difference from the historical average macronutrient consumption for each corresponding period of life. We predict that protein consumption should be reduced by half in early middle age and late middle age, whereas in late age it should increase. Our predictions are in line with results on humans and simpler organisms in the context of prolonging life.

[Alzheimer's disease - history of failures - a short opinion on the contemporary status of research]. / Choroba Alzheimera ­ historia niepowodzen - krótka opinia na temat wspólczesnego stanu badan.

Over 110 years have passed since first description of Alzheimer's disease (AD). Despite intensive research, AD remains incurable. The vast majority of clinical trials on new therapies aimed at suppressing undesirable aggregation of ß-amyloid have failed. This requires rethinking of the strategy to fight AD. A lot of evidence supports the maintenance of ß-amyloid as the central object of AD pathology, however, its aggregation into larger supramolecular structures is probably an intermediate stage and not the cause of the disease. In the early stages of life, cellular stresses lead to epigenetic changes resulting in later amyloidogenesis and subsequent neurodegeneration. This process takes decades of life trajectory. Therefore, possible therapies should take into account this fact and be applied early in the life and prolonged later. Diagnosis of epigenetic changes leading to AD should be a research priority. If this concept is true, it may give hope to reverse these changes.

Correction: The Calculator of Anti-Alzheimer's Diet. Macronutrients.

[This corrects the article DOI: 10.1371/journal.pone.0168385.].

The Calculator of Anti-Alzheimer's Diet. Macronutrients.

The opinions about optimal proportions of macronutrients in a healthy diet have changed significantly over the last century. At the same time nutritional sciences failed to provide strong evidence backing up any of the variety of views on macronutrient proportions. Herein we present an idea how these proportions can be calculated to find an optimal balance of macronutrients with respect to prevention of Alzheimer's Disease (AD) and dementia. These calculations are based on our published observation that per capita personal income (PCPI) in the USA correlates with age-adjusted death rates for AD (AADR). We have previously reported that PCPI through the period 1925-2005 correlated with AADR in 2005 in a remarkable, statistically significant oscillatory manner, as shown by changes in the correlation coefficient R (Roriginal). A question thus arises what caused the oscillatory behavior of Roriginal? What historical events in the life of 2005 AD victims had shaped their future with AD? Looking for the answers we found that, considering changes in the per capita availability of macronutrients in the USA in the period 1929-2005, we can mathematically explain the variability of Roriginal for each quarter of a human life. On the basis of multiple regression of Roriginal with regard to the availability of three macronutrients: carbohydrates, total fat, and protein, with or without alcohol, we propose seven equations (referred to as "the calculator" throughout the text) which allow calculating optimal changes in the proportions of macronutrients to reduce the risk of AD for each age group: youth, early middle age, late middle age and late age. The results obtained with the use of "the calculator" are grouped in a table (Table 4) of macronutrient proportions optimal for reducing the risk of AD in each age group through minimizing Rpredicted-i.e., minimizing the strength of correlation between PCPI and future AADR.

Correlation of Alzheimer's disease death rates with historical per capita personal income in the USA.

Alzheimer's disease (AD) is a progressive degenerating disease of complex etiology. A variety of risk factors contribute to the chance of developing AD. Lifestyle factors, such as physical, mental and social activity, education, and diet all affect the susceptibility to developing AD. These factors are in turn related to the level of personal income. Lower income usually coincides with lower level of education, lesser mental, leisure-social and physical activity, and poorer diet. In the present paper, we have analyzed the correlation of historical (1929-2011) per capita personal income (PCPI) for all states of the USA with corresponding age-adjusted AD death rates (AADR) for years 2000, 2005 and 2008. We found negative correlations in all cases, the highest one (R ≈ -0.65) for the PCPIs in the year 1970 correlated against the AADRs in 2005. From 1929 to 2005 the R value varies in an oscillatory manner, with the strongest correlations in 1929, 1970, 1990 and the weakest in 1950, 1980, 1998. Further analysis indicated that this oscillatory behavior of R is not artificially related to the economic factors but rather to delayed biological consequences associated with personal income. We conclude that the influence of the income level on the AD mortality in 2005 was the highest in the early years of life of the AD victims. Overall, the income had a significant, lifelong, albeit constantly decreasing, influence on the risk of developing AD. We postulate that the susceptibility of a population to late-onset AD (LOAD) is determined to a large extent by the history of income-related modifiable lifestyle risk factors. Among these risk factors, inappropriate diet has a significant contribution.

The ß-sheet breakers and π-stacking.

Fibrillation of ß-amyloid is recognized as a key process leading to the development of Alzheimer's disease. Small peptides called ß-sheet breakers were found to inhibit the process of ß-amyloid fibrillation and to dissolve amyloid fibrils in vitro, in vivo, and in cell culture studies [1,2]. The mechanism by which peptide inhibition takes place remains elusive and a detailed model needs to be established. Here, we present new insights into the possible role of consecutive Phe residues, present in the structure of ß-sheet breakers, supported by the results obtained by means of MD simulations. We performed a 30-ns MD of two ß-sheet breakers: iAß5 (LPFFD) and iAß6 (LPFFFD) which have two and three consecutive Phe residues, respectively. We have found that Phe rings in these peptides tend to form stacked conformations. For one of the peptides--iAß6--the calculated electrostatic contribution to free energy of one of the conformers with three rings stacked (c2) is significantly lower than that corresponding to the unstacked one (c1), two rings stacked (c0) and second conformer with three rings stacked (c3). This may favor the interaction of the c2 conformer with the target on amyloid fibril. We hypothesize that the mechanism of inhibition of amyloidogenesis by ß-sheet breaker involves competition among π-stacked Phe residues of the inhibitor and π-stacking within the ß-amyloid fibril. iAß6 may be a promising candidate for a lead compound of amyloidogenesis inhibitors.

[Selected aspects of amyloidogenesis]. / Wybrane aspekty amyloidogenezy.

Unwanted aggregation of proteins into amyloid fibrils caused by protein misfolding is connected with many human and animal diseases. Mechanism of this process is intensively studied. In the present paper main achievements in this field are described. Some researchers postulate that fibrils are the less harmful species and the most toxic species are the earlier aggregates on the path to mature fibrils..Therefore, monomeric, oligomeric and finally mature fibrils stages of the reaction of amyloidogenesis are reviewed with attention payed to the problem of structural polymorphism of fibrils.

Nature of cross-seeding barriers of amyloidogenesis.

The epidemics of bovine spongiform encephalopathy (BSE) several decades ago and present epidemics of chronic wasting disease (CWD) among cervids posed a threat of cross-species infections to humans or other animals. Therefore, the question as to the molecular nature of the species barriers to transmissibility of prion diseases is very important. We approached this problem theoretically, first developing a model of template-monomer interaction based on logical and topological grounds and on experimental data about cross-seeding of PrP 23-144 protein orthologs. Further, we propose that the strength of the cross-seeding barriers is proportional to dissimilarity of key amyloidogenic regions of the proteins. This dissimilarity can be measured by dissimilarity function we propose. Scaled on experimental data, this function predicts if cross-seeding can occur between different variants of PrP23-144. The resemblance of PrP23-144 cross-seeding barriers to the barriers of cross-species transmissibility of prion diseases is discussed. We suggest that a similar theoretical approach could be applied to predicting the occurrence of species barriers of prion diseases at least in part corresponding to the process of multiplication of infectious agent.

Direct interaction between prion protein and tubulin.

Recently published data show that the prion protein in its cellular form (PrP(C)) is a component of multimolecular complexes. In this report, zero-length cross-linking with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) allowed us to identify tubulin as one of the molecules interacting with PrP(C) in complexes observed in porcine brain extracts. We found that porcine brain tubulin added to these extracts can be cross-linked with PrP(C). Moreover, we observed that the 34 kDa species identified previously as full-length diglycosylated prion protein co-purifies with tubulin. Cross-linking of PrP(C) species separated by Cu(2+)-loaded immobilized metal affinity chromatography confirmed that only the full-length protein but not the N-terminally truncated form (C1) binds to tubulin. By means of EDC cross-linking and cosedimentation experiments, we also demonstrated a direct interaction of recombinant human PrP (rPrP) with tubulin. The stoichiometry of cosedimentation implies that rPrP molecules are able to bind both the alpha- and beta-isoforms of tubulin composing microtubule. Furthermore, prion protein exhibits higher affinity for microtubules than for unpolymerized tubulin.

Proteolytic processing and glycosylation influence formation of porcine prion protein complexes.

High level of heterogeneity seems to be a ubiquitous feature of mammalian PrPs (prion proteins) and may be relevant to the pathogenesis of prion diseases. In the present study, we describe the heterogeneity of PrP(C) (cellular form of PrP) from porcine brain. It was disclosed and characterized by a combination of one-dimensional PAGE and two-dimensional PAGE analyses with enzymic deglycosylation and copper-affinity experiments. We found that the identified two main populations of porcine PrP(C) consist of diglycosylated forms and correspond to the full-length (molecular mass 32-36 kDa) and proteolytically modified protein (molecular mass 25-30 kDa), known as C1. The two populations were fully separated during Cu2+-loaded immobilized metal affinity chromatography, indicating different affinity for copper ions. The more basic forms, migrating as species of higher molecular mass, exhibited stronger affinity for copper ions, whereas those with more acidic pI and of lower molecular mass were low-affinity Cu2+-binding molecules and thus could represent N-terminally truncated PrP(C). Size-exclusion chromatography revealed that most of the PrP(C) molecules in porcine brain extracts exist in the form of high-molecular-mass complexes (probably with other proteins). The heterogeneity of porcine PrP(C), resulting from proteolytic modification and glycosylation, influences its ability to assemble into these complexes. N-truncated molecules dominate over full-length PrP(C) in fractions of molecular mass over the range 65-130 kDa, whereas the full-length species are the major forms of PrP(C) present in the monomeric fraction and in complexes above 130 kDa. Two-dimensional PAGE analysis indicated that the complexed PrP(C) differs in the composition of pI forms from the monomers.

Ca2+ binding to myosin regulatory light chain affects the conformation of the N-terminus of essential light chain and its binding to actin.

We prepared a new type of skeletal myosin subfragment 1 (S1-MLC1F) containing both, the essential and the regulatory light chains, intact, by exchanging the essential light chains of papain S1 with bacterially expressed longer isoform (MLC1F) of this light chain. We then compared the enzymatic and structural properties of chymotryptic S1, papain S1, and S1-MLC1F in the presence and in the absence of Ca(2+) ions bound to the regulatory light chain. In the presence of Ca(2+), subfragment 1 containing both intact light chains exhibited lower V(max) and lower K(m) for actin activation of S1 ATPase. When S1-MLC1F was cross-linked to actin via the N-terminus of the essential light chain, the yield was much higher when Ca(2+) ions saturated the regulatory light chain. Limited proteolysis of the essential light chain in S1-MLC1F was significantly inhibited in the presence of calcium as compared to chymotryptic S1. We conclude that the effect of binding of Ca(2+) to the regulatory light chain is transmitted to the N-terminal extension of the longer isoform of the essential light chain. The resulting structure of the N-terminus is less susceptible to proteolytic digestion, binds tighter to actin, and has an inhibitory effect on actin-activated myosin ATPase. This new conformation of the N-terminus may be responsible for calcium induced myosin-linked modulation of striated muscle contraction.

The prion peptide forms ion channels in planar lipid bilayers.

One of the hypotheses concerning the pathogenic properties of the prion protein considers its influence on cellular ion homeostasis. Using the lipid bilayer technique, the influence of prion-derived peptides on the lipid bilayer conductance was characterized. To evaluate the physiological significance and possible pathological functions of the peptides, their effect on the membrane potential and respiration rate of hippocampal mitochondria was also studied. We used a peptide bearing the human prion protein sequence YSNQNNF (PrP [169-175]), and peptide SSQNNF (PrP [170-175]) bearing a naturally-occurring mutation in position 171 [N(r)S] linked to schizoaffective diseases in humans (Samaia, H.B., Mari, J.J., Vallada, H.P., Moura R.P., Simpson A.J.G., Brentani R.R. A prion-linked psychiatric disorder. Nature 390 (1997) 241). In this report, we show that PrP [170-175] N171S increases the conductance of planar lipid bilayers. Based on the conductance of single channel currents recorded in 500/500 mM KCl (cis/trans), we found a single channel conductance of 8 to 26 pS. The native prion peptide PrP [169-175] does not form ion channels in the lipid bilayer. Neither of the peptides significantly changed the membrane potential or respiration rate of isolated rat hippocampal mitochondria. We propose a possible mechanism for channel formation by aggregation of the prion-derived peptide.

The effects of the interaction of myosin essential light chain isoforms with actin in skeletal muscles.

In order to compare the ability of different isoforms of myosin essential light chain to interact with actin, the effect of the latter protein on the proteolytic susceptibility of myosin light chains (MLC-1S and MLC-1V - slow specific and same as ventricular isoform) from slow skeletal muscle was examined. Actin protects both slow muscle essential light chain isoforms from papain digestion, similarly as observed for fast skeletal muscle myosin (Nieznanska et al., 1998, Biochim. Biophys. Acta 1383: 71). The effect of actin decreases as ionic strength rises above physiological values for both fast and slow skeletal myosin, confirming the ionic character of the actin-essential light chain interaction. To better understand the role of this interaction, we examined the effect of synthetic peptides spanning the 10-amino-acid N-terminal sequences of myosin light chain 1 from fast skeletal muscle (MLC-1F) (MLCFpep: KKDVKKPAAA), MLC-1S (MLCSpep: KKDVPVKKPA) and MLC-1V (MLCVpep: KPEPKKDDAK) on the myofibrillar ATPase of fast and slow skeletal muscle. In the presence of MLCFpep, we observed an about 19% increase, and in the presence of MLCSpep about 36% increase, in the myofibrillar ATPase activity of fast muscle. On the other hand, in myofibrillar preparations from slow skeletal muscle, MLCSpep as well as MLCVpep caused a lowering of the ATPase activity by about 36%. The above results suggest that MLCSpep induces opposite effects on ATPase activity, depending on the type of myofibrils, but not through its specific N-terminal sequence - which differs from other MLC N-terminal peptides. Our observations lead to the conclusion that the action of different isoforms of long essential light chain is similar in slow and fast skeletal muscle. However the interaction of essential light chains with actin leads to different physiological effects probably depending on the isoforms of other myofibrillar proteins.