Significance The ubiquitin system controls a wide range of processes in cells and provides attractive drug targets for the treatment cancer other diseases. However, it has proved difficult to obtain inhibitors ligases that conjugate substrates, which there are hundreds. One class, HECT (homologous E6AP C terminus) domain ligases, receives from an E2 enzyme transfers substrate. We have selected bicyclic peptides block binding site individual as well small molecule, heclin (HECT ligase...

Ndfip1 and Ndfip2 are related endosomal membrane proteins that bind to activate members of the Nedd4 family E3 ubiquitin ligases. These ligases in turn affect receptor tyrosine kinase signaling by ubiquitinating several key components pathways. Here we investigate role Ndfip EGF signaling. We show they associate with PTEN, control ubiquitination abundance c-Cbl, Src kinases. Ndfip2, but not Ndfip1, also binds is phosphorylated Lyn, can act as a scaffold for phosphorylation potentially other...

Dishevelled is a pivot in Wnt signal transduction, controlling both β-catenin-dependent transcription to specify proliferative cell fates, and polarity other non-nuclear events post-mitotic cells. In response signals, or when present at high levels, forms signalosomes by dynamic polymerization. Its levels are controlled ubiquitylation, mediated various ubiquitin ligases, including NEDD4 family members that bind conserved PPxY motif (mammalian Dvl1-3). Here, we show Dvl2 binds the ligase WWP2...

We have isolated Spike, a novel and evolutionary conserved BH3-only protein. proteins constitute family of apoptosis inducers that mediate proapoptotic signals. In contrast to most this family, Spike was not found be associated with mitochondria. Furthermore, unlike the known proteins, could interact all tested Bcl-2 members, despite its BH3 domain being necessary for cell killing. Our findings indicate is localized endoplasmic reticulum. The reticulum an organelle has only recently been...

A genetic screen was established to clone apoptosis-inducing genes in a high-throughput format. It led the isolation of several proapoptotic whose proteins are localized mitochondria. One isolated is cytochrome b L (cybL also known as SDHC, C II-3 , or QPs-1), component respiratory chain complex II. further investigated because both cybL and another II, cybS, have recently been identified tumor suppressor proteins, some which act by controlling apoptosis. Our studies reveal that cell death...

Antibodies were raised against homogeneous preparations of component C the methylreductase system from Methanococcus voltae and Methanobacterium thermoautotrophicum . Cells these organisms fixed with paraformaldehyde and/or glutaraldehyde, sectioned, labeled antibodies colloidal gold-labeled protein A. In M. gold particles predominantly located in vicinity cytoplasmic membrane. rare cases a similar result was obtained also However, all but few ultrathin sections this bacterium, label...

Therapeutic strategies to provide local inhibition of mitogen mediated proliferation and migration human coronary artery smooth muscle cells (CASMC) by means drug-eluting stents have been shown enable effective limitation neointimal hyperplasia. However, currently available utilize compounds that may also adversely affect endothelial regrowth, thus possibly precipitating subsequent cardiac events. Accordingly, identification differentially inhibit cell could be substantial clinical...

Alpha-synuclein can form beta-sheet filaments, the accumulation of which plays a key role in development Parkinson's disease, dementia with Lewy bodies and multiple system atrophy. It has previously been shown that alpha-synuclein is substrate for HECT domain-containing ubiquitin ligase Nedd4, subject to ubiquitin-mediated endosomal degradation. We show here filaments are much better substrates ubiquitination vitro than monomeric alpha-synuclein, this increased susceptibility cannot be...

Activity-based probes (ABPs) have been used to dissect the biochemical/structural properties and cellular functions of deubiquitinases. However, their utility in studying cysteine-based E3 ubiquitin ligases has limited. In this study, we evaluate use ubiquitin-ABPs (Ub-VME Ub-PA) a novel set E2-Ub-ABPs on panel HECT ligases. Our vitro data show that can label domains. We also provide first evidence that, addition RBR ligase Parkin, catalytic domains NEDD4, UBE3C, HECTD1. Importantly,...

PCRctic is an innovative assay based on 16S rDNA PCR technology that has been designed to detect a single intact bacterium in specimen of cerebro-spinal fluid (CSF). The assay's potential for accurate, fast and inexpensive discrimination bacteria-free CSF makes it ideal adjunct confident exclusion bacterial meningitis newborn babies where the negative predictive value culture poor. This study aimed stress-test optimize "field conditions" attain clinically useful level specificity.The...

<h3></h3> Meningitis in newborn babies is difficult to rule out either clinically or microbiologically, which leads uncertainty with regards the initiation and length of antimicrobial treatment, risks under- over-treating. A different diagnostic approach that will offer high negative predictive value (NPV) needed overcome this problem. Such an shift emphasis on result resulting shorter antibiotic courses earlier discharges. <h3>Methods</h3> This novel molecular-biological assay has been...

<h3>Aims</h3> An assay based on 16S rDNA PCR technology has been designed to detect a single intact bacterium whilst eliminating free DNA from dead bacteria, thus offering unprecedented sensitivity and scope the analysis of bacterial carriage in clinical specimens. We hypothesised that application such an neonatal CSF will enable accurate, fast inexpensive discrimination bacteria-free specimens, have small but clinically acceptable false-positive rate. <h3>Methods</h3> Design PCRctic – novel...

Abstract Background: PCRctic is an innovative assay based on 16S rDNA PCR technology that has been designed to detect a single intact bacterium in specimen of cerebro-spinal fluid (CSF). The assay’s potential for accurate, fast and inexpensive discrimination bacteria-free CSF makes it ideal adjunct confident exclusion bacterial meningitis newborn babies where the negative predictive value culture poor. This study aimed stress-test optimize “field conditions” attain clinically useful level...

Abstract Background: PCRctic is an innovative assay based on 16S rDNA PCR technology that has been designed to detect a single intact bacterium in specimen of cerebro-spinal fluid (CSF). The assay’s potential for accurate, fast and inexpensive discrimination bacteria-free CSF makes it ideal adjunct confident exclusion bacterial meningitis newborn babies where the negative predictive value culture poor. This study aimed stress-test optimize “field conditions” attain clinically useful level...