Unraveling the mechanism of protein disaggregation through a ClpB-DnaK interaction.

HSP-100 protein machines, such as ClpB, play an essential role in reactivating protein aggregates that can otherwise be lethal to cells. Although the players involved are known, including the DnaK/DnaJ/GrpE chaperone system in bacteria, details of the molecular interactions are not well understood. Using methyl-transverse relaxation-optimized nuclear magnetic resonance spectroscopy, we present an atomic-resolution model for the ClpB-DnaK complex, which we verified by mutagenesis and functional assays.

A new perspective on Hsp104-mediated propagation and curing of the yeast prion [PSI (+) ].

Most prions in yeast form amyloid fibrils that must be severed by the protein disaggregase Hsp104 to be propagated and transmitted efficiently to newly formed buds. Only one yeast prion, [PSI (+) ], is cured by Hsp104 overexpression. We investigated the interaction between Hsp104 and Sup35, the priongenic protein in yeast that forms the [PSI (+) ] prion.

Insight into molecular basis of curing of [PSI+] prion by overexpression of 104-kDa heat shock protein (Hsp104).

Yeast prions are a powerful model for understanding the dynamics of protein aggregation associated with a number of human neurodegenerative disorders. The AAA+ protein disaggregase Hsp104 can sever the amyloid fibrils produced by yeast prions. This action results in the propagation of "seeds" that are transmitted to daughter cells during budding.

A multistage system for the automated detection of epileptic seizures in neonatal electroencephalography.

This paper describes the design and test results of a three-stage automated system for neonatal EEG seizure detection. Stage I of the system is the initial detection stage and identifies overlapping 5-second segments of suspected seizure activity in each EEG channel. In stage II, the detected segments from stage I are spatiotemporally clustered to produce multichannel candidate seizures.

Remodeling of protein aggregates by Hsp104.

Hsp104 is molecular chaperone in the AAA+ family of ATPases that specializes in the resolubilization and refolding of thermally denatured proteins in yeast. In addition to providing high levels of thermotolerance, Hsp104 plays a pivotal role in the propagation of yeast prions, self-replicating, amyloid-like aggregates that are inherited during mitosis and meiosis. In this review, the structure and function of Hsp104 is discussed, its functional interaction with other molecular chaperones, and a model for disaggregation and refolding is proposed.

The SmpB-tmRNA tagging system plays important roles in Streptomyces coelicolor growth and development.

The ssrA gene encodes tmRNA that, together with a specialized tmRNA-binding protein, SmpB, forms part of a ribonucleoprotein complex, provides a template for the resumption of translation elongation, subsequent termination and recycling of stalled ribosomes. In addition, the mRNA-like domain of tmRNA encodes a peptide that tags polypeptides derived from stalled ribosomes for degradation. Streptomyces are unique bacteria that undergo a developmental cycle culminating at sporulation that is at least partly controlled at the level of translation elongation by the abundance of a rare tRNA that decodes UUA codons found in a relatively small number of open reading frames prompting us to examine the role of tmRNA in S.

Peptide and protein binding in the axial channel of Hsp104. Insights into the mechanism of protein unfolding.

The AAA+ molecular chaperone Hsp104 mediates the extraction of proteins from aggregates by unfolding and threading them through its axial channel in an ATP-driven process. An Hsp104-binding peptide selected from solid phase arrays enhanced the refolding of a firefly luciferase-peptide fusion protein. Analysis of peptide binding using tryptophan fluorescence revealed two distinct binding sites, one in each AAA+ module of Hsp104.

The C-terminal extension of Saccharomyces cerevisiae Hsp104 plays a role in oligomer assembly.

The Saccharomyces cerevisiae protein Hsp104, a member of the Hsp100/Clp AAA+ family of ATPases, and its orthologues in plants (Hsp101) and bacteria (ClpB) function to disaggregate and refold thermally denatured proteins following heat shock and play important roles in thermotolerance. The primary sequences of fungal Hsp104's contain a largely acidic C-terminal extension not present in bacterial ClpB's. In this work, deletion mutants were used to determine the role this extension plays in Hsp104 structure and function.

Nucleocytoplasmic trafficking of the molecular chaperone Hsp104 in unstressed and heat-shocked cells.

Hsp104 is a molecular chaperone in yeast that restores solubility and activity to inactivated proteins after severe heat shock. We investigated the mechanisms that influence Hsp104 subcellular distribution in both unstressed and heat-shocked cells. In unstressed cells, Hsp104 and a green fluorescent protein-Hsp104 fusion protein were detected in both the nucleus and the cytoplasm.

Enhanced spatio-temporal clustering in the detection of neonatal seizures using context-based rules.

This work describes the clustering stage of a three-stage automated neonatal seizure detection system. This stage clusters spatio-temporally the short candidate seizure segments detected in prior stages, and then applies a variety of context-based rules to eliminate false detections and determine the final detected seizures. The work discusses important considerations in the implementation of rules and presents preliminary results.

Detection of pseudosinusoidal epileptic seizure segments in the neonatal EEG by cascading a rule-based algorithm with a neural network.

This paper presents an approach to detect epileptic seizure segments in the neonatal electroencephalogram (EEG) by characterizing the spectral features of the EEG waveform using a rule-based algorithm cascaded with a neural network. A rule-based algorithm screens out short segments of pseudosinusoidal EEG patterns as epileptic based on features in the power spectrum. The output of the rule-based algorithm is used to train and compare the performance of conventional feedforward neural networks and quantum neural networks.

BAG5 inhibits parkin and enhances dopaminergic neuron degeneration.

Loss-of-function mutations in the parkin gene, which encodes an E3 ubiquitin ligase, are the major cause of early-onset Parkinson's disease (PD). Decreases in parkin activity may also contribute to neurodegeneration in sporadic forms of PD. Here, we show that bcl-2-associated athanogene 5 (BAG5), a BAG family member, directly interacts with parkin and the chaperone Hsp70.

Saccharomyces cerevisiae Hsp104 enhances the chaperone capacity of human cells and inhibits heat stress-induced proapoptotic signaling.

Hsp104, the most potent thermotolerance factor in Saccharomyces cerevisiae, is an unusual molecular chaperone that is associated with the dispersal of aggregated, non-native proteins in vivo and in vitro. The close cooperation between Hsp100 oligomeric disaggregases and specific Hsp70 chaperone/cochaperone systems to refold and reactivate heat-damaged proteins has been dubbed a "bichaperone network". Interestingly, animal genomes do not encode a Hsp104 ortholog.

Amino acid substitutions in the C-terminal AAA+ module of Hsp104 prevent substrate recognition by disrupting oligomerization and cause high temperature inactivation.

Hsp104 is an important determinant of thermotolerance in yeast and is an unusual molecular chaperone that specializes in the remodeling of aggregated proteins. The structural requirements for Hsp104-substrate interactions remain unclear. Upon mild heat shock Hsp104 formed cytosolic foci in live cells that indicated co-localization of the chaperone with aggregates of thermally denatured proteins.

Evidence for an unfolding/threading mechanism for protein disaggregation by Saccharomyces cerevisiae Hsp104.

Saccharomyces cerevisiae Hsp104, a hexameric member of the Hsp100/Clp subfamily of AAA+ ATPases with two nucleotide binding domains (NBD1 and 2), refolds aggregated proteins in conjunction with Hsp70 molecular chaperones. Hsp104 may act as a "molecular crowbar" to pry aggregates apart and/or may extract proteins from aggregates by unfolding and threading them through the axial channel of the Hsp104 hexamer. Targeting Tyr-662, located in a Gly-Tyr-Val-Gly motif that forms part of the axial channel loop in NBD2, we created conservative (Phe and Trp) and non-conservative (Ala and Lys) amino acid substitutions.