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W Baumeister

Publications and source records attributed to W Baumeister.

At least 19 recordsLinked to original sources
PubMed1999

Structure of VAT, a CDC48/p97 ATPase homologue from the archaeon Thermoplasma acidophilum as studied by electron tomography.

Valosine-containing protein-like ATPase from Thermoplasma acidophilum is a member of the superfamily of ATPases associated with a diversity of cellular activities and is closely related to CDC48 from yeast and p97 from higher eukaryotes and more distantly to N-ethylmaleimide-sensitive fusion protein. We have used electron tomography to obtain low-resolution (2-2.5 nm) three-dimensional maps of both the whole 500 kDa complex and the N-terminally truncated valosine-containing protein-like ATPase from T. acidophilum complex lacking the putative substrate binding domain.

Adenosine Triphosphatases↗
PubMed1999

A giant protease with potential to substitute for some functions of the proteasome.

An alanyl-alanyl-phenylalanyl-7-amino-4-methylcoumarin-hydrolyzing protease particle copurifying with 26S proteasomes was isolated and identified as tripeptidyl peptidase II (TPPII), a cytosolic subtilisin-like peptidase of unknown function. The particle is larger than the 26S proteasome and has a rod-shaped, dynamic supramolecular structure. TPPII exhibits enhanced activity in proteasome inhibitor-adapted cells and degrades polypeptides by exo- as well as predominantly trypsin-like endoproteolytic cleavage. TPPII may thus participate in extralysosomal polypeptide degradation and may in part account for nonproteasomal epitope generation as postulated for certain major histocompatibility complex class I alleles. In addition, TPPII may be able to substitute for some metabolic functions of the proteasome.

Acetylcysteine↗
PubMed1999

Proteasomes and other self-compartmentalizing proteases in prokaryotes.

The proteasome represents the major non-lysosomal proteolytic system in eukaryotes. It confines proteolytic activity to an inner compartment that is accessible to unfolded proteins only. The strategy of controlling intracellular breakdown of proteins by self-compartmentalization is also used by different types of prokaryotic energy-dependent proteases. Genomic sequencing data reveal that various combinations of these energy-dependent proteases occur in prokaryotic cells from different lineages.

Adenosine Triphosphatases↗
PubMed1999

Functional analysis of the proteasome regulatory particle.

We have developed S. cerevisiae as a model system for mechanistic studies of the 26S proteasome. The subunits of the yeast 19S complex, or regulatory particle (RP), have been defined, and are closely related to those of mammalian proteasomes. The multiubiquitin chain binding subunit (S5a/Mcb1/Rpn10) was found, surprisingly, to be nonessential for the degradation of a variety of ubiquitin-protein conjugates in vivo. Biochemical studies of proteasomes from deltarpn10 mutants revealed the existence of two structural subassemblies within the RP, the lid and the base. The lid and the base are both composed of 8 subunits. By electron microscopy, the base and the lid correspond to the proximal and distal masses of the RP, respectively. The base is sufficient to activate the 20S core particle for degradation of peptides, but the lid is required for ubiquitin-dependent degradation. The lid subunits share sequence motifs with components of the COP9/signalosome complex, suggesting that these functionally diverse particles have a common evolutionary ancestry. Analysis of equivalent point mutations in the six ATPases of the base indicate that they have well-differentiated functions. In particular, mutations in one ATPase gene, RPT2, result in an unexpected defect in peptide hydrolysis by the core particle. One interpretation of this result is that Rpt2 participates in gating of the channel through which substrates enter the core particle.

Adenosine Triphosphatases↗
PubMed1999

[Genotoxicity of stack gas condensates of Bavarian waste incineration plants. II. Suitability of fast bacterial tests of emission monitoring].

The genotoxicity of stack gas condensates of 21 waste incineration plants (located in Bavaria) was examined in the years 1990-1995 using two bacterial short time tests. The SOS chromotest was carried out with the tester strains Escherichia coli PQ37, PQ243 and PQ300. In addition, for the purpose of comparison, the Ames-Test was performed for selected examples with the tester strains Salmonella typhimurium TA97, TA98, TA100 and TA1537. In a pilot study, carried out in the years 1990 to 1991, the stack gas condensates from five plants were examined. They showed clear genotoxic and mutagenic effects. On the other hand, in subsequent tests we generally discovered only weak inductions for 9 of 18 crude and 24 of 78 clean gas condensate extracts, mostly after metabolic activation. Four plants were tested continuously in the years 1992 to 1995. Three of them showed a clear reduction of the detectable genotoxic potential. The fourth one gave negligible SOS inducing emissions in the whole examining period. On the other hand, for 6 of the 21 tested plants we found chromotest positive results even at the last test point. Correlations between the SOS inducing potential of the stack gas condensates and the analytical parameters detected at the same time (6 summary parameters, 24 inorganic and 63 organic chemical parameters) were not evident. Only the two highest emissions of nitropyrenes were associated with SOS inductions. Organic substances which are not analytically detected or synergistic effects might be responsible for the SOS inducing potency of the other genotoxic stack gas condensates.

Air Pollutants↗
PubMed1998

Late events in the assembly of 20S proteasomes.

Electron microscopy and STEM mass measurements have been used to characterize late intermediates in the assembly pathway of wildtype and mutant Rhodococcus proteasomes. A proteolytically inactive and processing-incompetent mutant, betaK33A, allowed a short-lived late intermediate of the pathway to be captured, the preholoproteasome. In this fully assembled 20S complex the 14 propeptides with an aggregate mass of 100 kDa fill the whole central cavity and most of the two antechambers. It is further shown that in wildtype Rhodococcus proteasomes the propeptides are degraded in a processive manner undergoing multiple cleavages before the products are discharged and the inner cavities are cleared. It appears that the docking of two half-proteasomes, i.e., preholoproteasome formation, is sufficient to trigger autocleavage of the Gly-1/Thr1 bond necessary for active site formation and the subsequent degradation of the propeptides.

Amino Acid Substitution↗
PubMed1998

The role of tricorn protease and its aminopeptidase-interacting factors in cellular protein degradation.

Tricorn protease was previously described as the core enzyme of a modular proteolytic system displaying multicatalytic activity. Here we elucidate the mode of cooperation between Tricorn and its interacting factors, and we identify two additional factors, F2 and F3, closely related aminopeptidases of 89 kDa. In conjunction with these three factors, Tricorn degrades oligopeptides in a sequential manner, yielding free amino acids. We have been able to reconstitute a proteolytic pathway comprising the proteasome, Tricorn, and its interacting factors, F1, F2, and F3, which converts proteins efficiently into amino acids. Therefore, it is quite likely that Tricorn also acts in vivo downstream of the proteasome and, in cooperation with its interacting factors, completes protein catabolic pathways.

Amino Acid Sequence↗
PubMed1998

The junctional pore complex, a prokaryotic secretion organelle, is the molecular motor underlying gliding motility in cyanobacteria.

BACKGROUND: Whereas most bacteria move by means of flagella, some prokaryotes move by gliding. In cyanobacteria, gliding motility is a slow uniform motion which is invariably accompanied by a continuous secretion of slime. On the basis of these characteristics, a model has been proposed in which the gliding motility of cyanobacteria depends on the steady secretion of slime using specific pores, as well as the interaction of the slime with the filament surface and the underlying substrate. RESULTS: The structures of the pore apparatus of two different filamentous cyanobacteria have been characterized. In both species, pores are formed by a hitherto uncharacterized type of prokaryotic organelle that spans the entire multilayered cell wall and possesses structural properties expected for an organelle that is involved in the rapid secretion of extracellular carbohydrates. Light microscopic observations of the secretion process provided direct evidence that the pore complexes are the actual sites of slime secretion, that the secreted slime fibrils are elongated at about the same rate as the filament glides (up to 3 micrometer s-1), and that gliding movements are caused directly by the secretion of slime. CONCLUSIONS: It has been known for a long time that carbohydrate secretion has an important role in the gliding motility of various prokaryotes. Our results strongly suggest that slime secretion is not only a prerequisite for this peculiar type of motility in cyanobacteria, but also directly generates the necessary thrust for locomotion.

Anabaena↗
PubMed1998

Decelerated degradation of short peptides by the 20S proteasome.

Based on a twelve residue master peptide comprising all five specific cleavage sites defined for the proteasome, a set of variant peptides was generated in order to probe specificity and to elucidate the mechanism which determines product size. It is shown that the rate of degradation by the 20S proteasome from Thermoplasma acidophilum depends critically on the length of the peptide substrate. Peptides of 14 residues and longer are degraded much faster than shorter peptides although the sites of cleavage remain unchanged. The decelerated degradation of peptides shorter than 14 residues explains the accumulation of products with an average length of seven to nine residues.

Amino Acid Sequence↗
PubMed1998

A subcomplex of the proteasome regulatory particle required for ubiquitin-conjugate degradation and related to the COP9-signalosome and eIF3.

The proteasome consists of a 20S proteolytic core particle (CP) and a 19S regulatory particle (RP), which selects ubiquitinated substrates for translocation into the CP. An eight-subunit subcomplex of the RP, the lid, can be dissociated from proteasomes prepared from a deletion mutant for Rpn10, an RP subunit. A second subcomplex, the base, contains all six proteasomal ATPases and links the RP to the CP. The base is sufficient to activate the CP for degradation of peptides or a nonubiquitinated protein, whereas the lid is required for ubiquitin-dependent degradation. By electron microscopy, the base and the lid correspond to the proximal and distal masses of the RP, respectively. The lid subunits share sequence motifs with components of the COP9/signalosome complex and eIF3, suggesting that these functionally diverse particles have a common evolutionary ancestry.

COP9 Signalosome Complex↗
PubMed1998

The thermosome: archetype of group II chaperonins.

The thermosome, the chaperonin of the archaea, and its homologue from the cytosol of eukaryotes, known as TRiC or CCT, form a distinct subfamily of the chaperonins that does not depend on a co-chaperonin for protein folding activity. Recent structural data obtained by cryo- electron microscopy and X-ray crystallography provide the first insights into a novel mechanism remarkably different from that of the bacterial GroEL-GroES system.

Animals↗
PubMed1998

Characterization of ARC, a divergent member of the AAA ATPase family from Rhodococcus erythropolis.

A gene encoding a AAA ATPase was discovered in the 5' region of the second operon of 20 S proteasome subunits in the nocardioform actinomycete Rhodococcus erythropolis NI86/21. The gene was cloned and expressed in Escherichia coli. The protein, ARC (AAA ATPase forming Ring-shaped Complexes), is a divergent member of the AAA family. The deduced product of the arc gene is 591 residues long (66 kDa). The purified protein possesses a low, N-ethylmaleimide-sensitive ATPase activity and forms rings of six subunits, arranged symmetrically around a central opening or cavity. Two-dimensional crystals grown on lipid monolayers yielded images of the ATPase molecules in "end-on" orientation at 1.9 nm resolution.

Amino Acid Sequence↗
PubMed1998

Proteasome function is dispensable under normal but not under heat shock conditions in Thermoplasma acidophilum.

Hitherto the biology of proteolysis in prokaryotes, particularly in archaea, is only poorly understood. We have used the tri-peptide vinyl sulfone inhibitor carboxybenzyl-leucyl-leucyl-leucine vinyl sulfone (Z-L3VS) to study the in vivo function of proteasomes in Thermoplasma acidophilum. Z-L3VS is a potent inhibitor of the Thermoplasma proteasome and is capable of modifying 75 to 80% of the proteasomal beta-subunits in cell cultures. Inhibition of proteasomes has only marginal effects under normal growth conditions. Under heat shock conditions, however, the effects of proteasome inhibition are much more severe, to the extent of complete cell growth arrest. These data suggest that other proteolytic systems may exist that can compensate for the loss of proteasome function in T. acidophilum.

Cell Division↗
PubMed1998

26S proteasome structure revealed by three-dimensional electron microscopy.

In 26S proteasomes, "19S cap complexes" associate with either one or both ends of the barrel-shaped 20S core complex. These regulatory complexes which comprise about 20 different subunits, including 6 ATPases of the AAA family, are thought to recognize ubiquitinated substrate proteins, to dissociate and unfold them before threading them into the 20S core where they are degraded. Here, we examine the structure of 26S proteasomes from Drosophila embryos and Xenopus oocytes by electron microscopy. Image analysis reveals a rather flexible linkage between the 19S caps and the 20S core, with a peculiar wagging-type movement of the caps relative to the core. At this stage of the analysis, it is not clear whether this movement is relevant in terms of function. Three-dimensional reconstructions, taking this into account, provide first insights into the remarkably complex structure of the 19S caps and allows us to put forward a composite model of the entire 26S complex.

Animals↗
PubMed1998

Electron tomography of ice-embedded prokaryotic cells.

Whole cells of archaea were embedded in vitreous ice by plunge freezing and investigated by automated energy-filtered electron tomography at 120 kV. The embedded cells were between 300 and 750 nm thick, and their structures were reconstructed to a resolution of 20-40 nm from tilt series comprising 50-140 images. The dose was kept within tolerable limits. A resolution of 20 nm allowed visualization of the individual stalks of the S-layer of Pyrobaculum aerophilum cells, which had undergone partial lysis, in three dimensions. The attainable resolution for low-dose electron tomography under different experimental conditions was theoretically investigated in terms of the specimen thickness. To obtain 2-nm resolution at 120 kV (300 kV), the specimen must not be thicker than 100 nm (150 nm). For a resolution of 10 nm, the maximum thickness is 450 nm (700 nm). An accelerating voltage of 300 kV is advantageous, mainly for specimens thicker than 100 nm. Experimental investigations so far have resulted in a resolution that is worse by a factor of 2-5 as compared to theory.

Automation↗