Clustering and Sharing Incentives in BitTorrent Systems

Peer-to-peer protocols play an increasingly instrumental role in Internet content distribution. Consequently, it is important to gain a full understanding of how these protocols behave in practice and how their parameters impact overall performance. We present the first experimental investigation of the peer selection strategy of the popular BitTorrent protocol in an instrumented private torrent. By observing the decisions of more than 40 nodes, we validate three BitTorrent properties that, though widely believed to hold, have not been demonstrated experimentally. These include the clustering of similar-bandwidth peers, the effectiveness of BitTorrent's sharing incentives, and the peers' high average upload utilization. In addition, our results show that BitTorrent's new choking algorithm in seed state provides uniform service to all peers, and that an underprovisioned initial seed leads to the absence of peer clustering and less effective sharing incentives. Based on our observations, we provide guidelines for seed provisioning by content providers, and discuss a tracker protocol extension that addresses an identified limitation of the protocol

Moment-Generating Algorithm for Response Time in Processor Sharing Queueing Systems

Response times are arguably the most representative and important metric for measuring the performance of modern computer systems. Further, service level agreements (SLAs), ranging from data centres to smartphone users, demand quick and, equally important, predictable response times. Hence, it is necessary to calculate moments, at least, and ideally response time distributions, which is not straightforward. A new moment-generating algorithm for calculating response times analytically is obtained, based on M/M/1 processor sharing (PS) queueing models. This algorithm is compared against existing work on response times in M/M/1-PS queues and extended to M/M/1 discriminatory PS queues. Two real-world case studies are evaluated

Dynamics of the Acetylcholinesterase Tetramer

Acetylcholinesterase rapidly hydrolyzes the neurotransmitter acetylcholine in cholinergic synapses, including the neuromuscular junction. The tetramer is the most important functional form of the enzyme. Two low-resolution crystal structures have been solved. One is compact with two of its four peripheral anionic sites (PAS) sterically blocked by complementary subunits. The other is a loose tetramer with all four subunits accessible to solvent. These structures lacked the C-terminal amphipathic t-peptide (WAT domain) that interacts with the proline-rich attachment domain (PRAD). A complete tetramer model (AChEt) was built based on the structure of the PRAD/WAT complex and the compact tetramer. Normal mode analysis suggested that AChEt could exist in several conformations with subunits fluctuating relative to one another. Here, a multiscale simulation involving all-atom molecular dynamics and Cα-based coarse-grained Brownian dynamics simulations was carried out to investigate the large-scale intersubunit dynamics in AChEt. We sampled the ns-μs timescale motions and found that the tetramer indeed constitutes a dynamic assembly of monomers. The intersubunit fluctuation is correlated with the occlusion of the PAS. Such motions of the subunits “gate” ligand-protein association. The gates are open more than 80% of the time on average, which suggests a small reduction in ligand-protein binding. Despite the limitations in the starting model and approximations inherent in coarse graining, these results are consistent with experiments which suggest that binding of a substrate to the PAS is only somewhat hindered by the association of the subunits

Heterologous Amyloid Seeding: Revisiting the Role of Acetylcholinesterase in Alzheimer's Disease

Neurodegenerative diseases associated with abnormal protein folding and ordered aggregation require an initial trigger which may be infectious, inherited, post-inflammatory or idiopathic. Proteolytic cleavage to generate vulnerable precursors, such as amyloid-β peptide (Aβ) production via β and γ secretases in Alzheimer's Disease (AD), is one such trigger, but the proteolytic removal of these fragments is also aetiologically important. The levels of Aβ in the central nervous system are regulated by several catabolic proteases, including insulysin (IDE) and neprilysin (NEP). The known association of human acetylcholinesterase (hAChE) with pathological aggregates in AD together with its ability to increase Aβ fibrilization prompted us to search for proteolytic triggers that could enhance this process. The hAChE C-terminal domain (T40, AChE575-614) is an exposed amphiphilic α-helix involved in enzyme oligomerisation, but it also contains a conformational switch region (CSR) with high propensity for conversion to non-native (hidden) β-strand, a property associated with amyloidogenicity. A synthetic peptide (AChE586-599) encompassing the CSR region shares homology with Aβ and forms β-sheet amyloid fibrils. We investigated the influence of IDE and NEP proteolysis on the formation and degradation of relevant hAChE β-sheet species. By combining reverse-phase HPLC and mass spectrometry, we established that the enzyme digestion profiles on T40 versus AChE586-599, or versus Aβ, differed. Moreover, IDE digestion of T40 triggered the conformational switch from α- to β-structures, resulting in surfactant CSR species that self-assembled into amyloid fibril precursors (oligomers). Crucially, these CSR species significantly increased Aβ fibril formation both by seeding the energetically unfavorable formation of amyloid nuclei and by enhancing the rate of amyloid elongation. Hence, these results may offer an explanation for observations that implicate hAChE in the extent of Aβ deposition in the brain. Furthermore, this process of heterologous amyloid seeding by a proteolytic fragment from another protein may represent a previously underestimated pathological trigger, implying that the abundance of the major amyloidogenic species (Aβ in AD, for example) may not be the only important factor in neurodegeneration

Pro-apoptotic protein–protein interactions of the extended N-AChE terminus

The N-terminally extended “synaptic” acetylcholinesterase variant N-AChE-S operates to promote apoptosis; however, the protein partners involved in this function remain unknown. Here, we report that when microinjected to fertilized mouse oocytes, N-AChE-S caused embryonic death as early as the zygotic stage. To identify the putative protein partners involved, we first tried yeast two hybrid screening, but this approach failed, probably because of the N-AChE-S-induced lethality. In contrast, sequence analysis and a corresponding peptide array revealed possible partners, which were validated by co-immunoprecipitation. These include the kinases GSK3, Aurora and GAK, the membrane integrin receptors, and the death receptor FAS. Each of these could potentially modulate N-AChE-S-induced apoptosis with possible therapeutic value for the treatment of Alzheimer’s disease

N-Acetylcholinesterase-Induced Apoptosis in Alzheimer's Disease

Background: Alzheimer’s disease (AD) involves loss of cholinergic neurons and Tau protein hyper-phosphorylation. Here, we report that overexpression of an N-terminally extended ‘‘synaptic’ ’ acetylcholinesterase variant, N-AChE-S is causally involved in both these phenomena. Methodology and Principal Findings: In transfected primary brain cultures, N-AChE-S induced cell death, morphological impairments and caspase 3 activation. Rapid internalization of fluorescently labeled fasciculin-2 to N-AChE-S transfected cells indicated membranal localization. In cultured cell lines, N-AChE-S transfection activated the Tau kinase GSK3, induced Tau hyper-phosphorylation and caused apoptosis. N-AChE-S-induced cell death was suppressible by inhibiting GSK3 or caspases, by enforced overexpression of the anti-apoptotic Bcl2 proteins, or by AChE inhibition or silencing. Moreover, inherent N-AChE-S was upregulated by stressors inducing protein misfolding and calcium imbalances, both characteristic of AD; and in cortical tissues from AD patients, N-AChE-S overexpression coincides with Tau hyper-phosphorylation. Conclusions: Together, these findings attribute an apoptogenic role to N-AChE-S and outline a potential value to ACh

A Soluble Acetylcholinesterase Provides Chemical Defense against Xenobiotics in the Pinewood Nematode

The pinewood nematode genome encodes at least three distinct acetylcholinesterases (AChEs). To understand physiological roles of the three pinewood nematode AChEs (BxACE-1, BxACE-2, and BxACE-3), BxACE-3 in particular, their tissue distribution and inhibition profiles were investigated. Immunohistochemistry revealed that BxACE-1 and BxACE-2 were distributed in neuronal tissues. In contrast, BxACE-3 was detected from some specific tissues and extracted without the aid of detergent, suggesting its soluble nature unlike BxACE-1 and BxACE-2. When present together, BxAChE3 significantly reduced the inhibition of BxACE-1 and BxACE-2 by cholinesterase inhibitors. Knockdown of BxACE-3 by RNA interference significantly increased the toxicity of three nematicidal compounds, supporting the protective role of BxACE-3 against chemicals. In summary, BxACE-3 appears to have a non-neuronal function of chemical defense whereas both BxACE-1 and BxACE-2 have classical neuronal function of synaptic transmission

Acetilkolinesteraza u eritrocitima i butirilkolinesteraza u plazmi - Važni pokazatelji za liječenje osoba otrovanih organofosfornim spojevima

Inhibition of acetylcholinesterase (AChE) is regarded as the primary toxic mechanism of organophosphorus compounds (OP). Therapeutic strategies are directed to antagonise overstimulation of muscarinic receptors with atropine and to reactivate inhibited AChE with oximes. Reactivation is crucial within the neuromuscular synapse, where atropine is ineffective, since peripheral neuromuscular block eventually leads to respiratory failure. Patients with OP intoxication have to be identified as early as possible. During an international NBC-defence exercise anesthetised pigs were poisoned with sarin, followed by treatment with atropine and oxime. Blood samples were drawn and red blood cell (RBC)-AChE activity determined with a fielded test system on-site. Within a few minutes the poisoning was verified. After administration of HI-6, RBC-AChE activity increased rapidly. Blood samples were reanalysed in our laboratory in Munich. Almost identical course of the AChE activities was recorded by both systems. The more comprehensive cholinesterase status was determined in Munich. Oxime administration can be stopped when AChE is aged completely, but has to be continued as long as poison is present in the body and reactivation is possible. To aid the on-site physician in optimising diagnosis and treatment, a fielded test system should be available to allow rapid determination of the complete cholinesterase status.Inhibicija acetilkolinesteraze (AChE) smatra se primarnim mehanizmom toksičnoga djelovanja organofosfornih spojeva (OP). Strategije liječenja idu za zaustavljanjem prekomjerne stimulacije muskarinskih receptora atropinom i reaktiviranjem inhibiranog AChE oksimima. Ključna je reaktivacija u neuromuskularnoj sinapsi, u kojoj atropin nije djelotvoran, budući da neuromuskularna blokada u konačnici vodi do prestanka disanja. Važno je što ranije prepoznati otrovanje organofosfornim spojem. U jednoj međunarodnoj vježbi zaštite od nuklearnog, biološkog i kemijskog napada svinje pod anestezijom otrovane su sarinom te liječene atropinom i oksimom. Uzeti su im uzorci krvi te s pomoću terenskoga testa na licu mjesta određena aktivnost AChE u eritrocitima. Otrovanje je potvrđeno za nekoliko minuta. Nakon primjene HI-6, aktivnost AChE brzo je porasla. Isti su uzorci krvi ponovno analizirani u našem laboratoriju u Münchenu. Oba su testa zabilježila gotovo istovjetan tijek aktivnosti AChE. U Münchenu je međutim napravljen potpuniji nalaz kolinesteraza. Liječenje oksimima može se prekinuti kada AChE potpuno “ostari” (tj. dealkilira), ali ga valja nastaviti dokle god je otrov u tijelu, a reaktivacija moguća. Liječnici na terenu trebali bi raspolagati terenskim testovima radi brzoga i potpunog utvrđivanja statusa kolinesteraza, a time i kvalitetnije dijagnoze

Cholinesterases: Structure, Role, and Inhibition

Acetilkolinesteraza (AChE; E.C. 3.1.1.7) i butirilkolinesteraza (BChE; E.C. 3.1.1.8) enzimi su koji se zbog svoje uloge u organizmu intenzivno istražuju unutar područja biomedicine i toksikologije. Iako strukturno homologni, ovi enzimi razlikuju se prema katalitičkoj aktivnosti, odnosno specifi čnosti prema supstratima koje mogu hidrolizirati te selektivnosti za vezanje mnogih liganada. U ovom radu dan je pregled dosadašnjih istraživanja kolinesteraza i njihovih interakcija s ligandima i inhibitorima te su izdvojene aminokiseline aktivnog mjesta koje sudjeluju u tim interakcijama.Enzymes acetylcholinesterase (AChE; E.C. 3.1.1.7) and butyrylcholinesterase (BChE; E.C. 3.1.1.8) have intensively been investigated in biomedicine and toxicology due to important role in organisms. Even if structurally homologous, they differ in catalytic activity, specificity, for substrates, and selectivity in binding to many ligands. This paper compiles the results of research on cholinesterases and their interactions with ligands and inhibitors, and identifies amino acids of active sites involved in these interactions