Biomedical subjects

Sonja-Verena Albers

Publications and source records attributed to Sonja-Verena Albers.

11 recordsLinked to original sources

PubMed2006

Thermodynamics of the ATPase cycle of GlcV, the nucleotide-binding domain of the glucose ABC transporter of sulfolobus solfataricus.

ATP-binding cassette transporters drive the transport of substrates across the membrane by the hydrolysis of ATP. They typically have a conserved domain structure with two membrane-spanning domains that form the transport channel and two cytosolic nucleotide-binding domains (NBDs) that energize the transport reaction. Binding of ATP to the NBD monomer results in formation of a NBD dimer. Hydrolysis of the ATP drives the dissociation of the dimer. The thermodynamics of distinct steps in the ATPase cycle of GlcV, the NBD of the glucose ABC transporter of the extreme thermoacidophile Sulfolobus solfataricus, were studied by isothermal titration calorimetry using the wild-type protein and two mutants, which are arrested at different steps in the ATP hydrolytic cycle. The G144A mutant is unable to dimerize, while the E166A mutant is defective in dimer dissociation. The ATP, ADP, and AMP-PNP binding affinities, stoichiometries, and enthalpies of binding were determined at different temperatures. From these data, the thermodynamic parameters of nucleotide binding, NBD dimerization, and ATP hydrolysis were calculated. The data demonstrate that the ATP hydrolysis cycle of isolated NBDs consists of consecutive steps where only the final step of ADP release is energetically unfavorable.

ATP-Binding Cassette Transporters↗

PubMed2006

Protein secretion in the Archaea: multiple paths towards a unique cell surface.

Archaea are similar to other prokaryotes in most aspects of cell structure but are unique with respect to the lipid composition of the cytoplasmic membrane and the structure of the cell surface. Membranes of archaea are composed of glycerol-ether lipids instead of glycerol-ester lipids and are based on isoprenoid side chains, whereas the cell walls are formed by surface-layer proteins. The unique cell surface of archaea requires distinct solutions to the problem of how proteins cross this barrier to be either secreted into the medium or assembled as appendages at the cell surface.

Archaea↗

PubMed2006

Regulation of expression of the arabinose and glucose transporter genes in the thermophilic archaeon Sulfolobus solfataricus.

Sugar uptake in Sulfolobus solfataricus, a thermoacidophilic archaeon, occurs through high-affinity binding of protein-dependent ABC transporters. We have investigated the expression patterns of two sugar transport operons, that is, the glucose and arabinose transporters. Analysis of the araS promoter activity, and the mRNA and protein levels in S. solfataricus cells grown on different carbon sources showed that expression of the arabinose transporter gene cluster is highly regulated and dependent on the presence of arabinose in the medium. Glucose in the growth medium repressed the expression of the arabinose transport genes. By means of primer extension, the transcriptional start site for the arabinose operon was mapped. Interestingly, expression of the arabinose transporter is down-regulated by addition of a selective set of amino acids to the medium. Expression of the glucose transporter genes appeared constitutive. These data confirm the earlier observation of a catabolite repression-like system in S. solfataricus.

Amino Acids↗

PubMed2006

Active-site residues in the type IV prepilin peptidase homologue PibD from the archaeon Sulfolobus solfataricus.

Archaeal preflagellin peptidases and bacterial type IV prepilin peptidases belong to a family of aspartic acid proteases that cleave the leader peptides of precursor proteins with type IV prepilin signal sequences. The substrate repertoire of PibD from the crenarchaeon Sulfolobus solfataricus is unusually diverse. In addition to flagellin, PibD cleaves three sugar-binding proteins unique to this species and a number of proteins with unknown function. Here we demonstrate that PibD contains two aspartic acid residues that are essential for cleavage activity. An additional pair of aspartic acids in a large cytoplasmic loop is also important for function and is possibly involved in leader peptide recognition. Combining the results of transmembrane segment predictions and cysteine-labeling experiments, we suggest a membrane topology model for PibD with the active-site aspartic acid residues exposed to the cytosol.

Archaeal Proteins↗

PubMed2005

Analysis of ATPases of putative secretion operons in the thermoacidophilic archaeon Sulfolobus solfataricus.

Gram-negative bacteria use a wide variety of complex mechanisms to secrete proteins across their membranes or to assemble secreted proteins into surface structures. As most archaea only possess a cytoplasmic membrane surrounded by a membrane-anchored S-layer, the organization of such complexes might be significantly different from that in Gram-negative bacteria. Five proteins of Sulfolobus solfataricus, SSO0120, SSO0572, SSO2316, SSO2387 and SSO2680, which are homologous to secretion ATPases of bacterial type II, type IV secretion systems and the type IV pili assembly machinery, were identified. The operon structures of these putative secretion systems encoding gene clusters and the expression patterns of the ATPases under different growth conditions were determined, and it was established that all five putative ATPases do show a divalent cation-dependent ATPase activity at high temperature. These results show that the archaeal secretion systems are related to the bacterial secretion systems and might be powered in a similar way.

Adenosine Triphosphatases↗

PubMed2004

Insights into ABC transport in archaea.

In archaea, ATP-binding cassette (ABC) transporters play a crucial role in substrate uptake, export, and osmoregulation. Archael substrate-binding-protein-dependent ABC transporters are equipped with a very high affinity for their cognate substrates which provide these organisms with the ability to efficiently scavenge substrates from their environment even when present only at low concentration. Further adaptations to the archaeal way of life are especially found in the domain organization and anchoring of the substrate-binding proteins to the membrane. Examination of the signal peptides of binding proteins of 14 archael genomes showed clear differences between euryarchaeotes and crenarchaeotes. Furthermore, a profiling and comparison of ABC transporters in the three sequenced pyrococcal strains was performed.

ATP-Binding Cassette Transporters↗

PubMed2003

Formation of the productive ATP-Mg2+-bound dimer of GlcV, an ABC-ATPase from Sulfolobus solfataricus.

The ABC-ATPase GlcV from Sulfolobus solfataricus energizes an ABC transporter mediating glucose uptake. In ABC transporters, two ABC-ATPases are believed to form a head-to-tail dimer, with both monomers contributing conserved residues to each of the two productive active sites. In contrast, isolated GlcV, although active, behaves apparently as a monomer in the presence of ATP-Mg(2+), AMPPNP-Mg(2+) or ATP alone. To resolve the oligomeric state of the active form of GlcV, we analysed the effects of changing the putative catalytic base, residue E166, into glutamine or alanine. Both mutants are, to different extents, defective in ATP hydrolysis, and gel-filtration experiments revealed their dimerization in the presence of ATP-Mg(2+). Mutant E166Q forms dimers also in the presence of ATP alone, without Mg(2+), whereas dimerization of mutant E166A requires both ATP and Mg(2+). These results confirm earlier reports for other ABC-ATPases, but for the first time suggest the occurrence of a fast equilibrium between ATP-bound monomers and ATP-bound dimers. We further mutated two highly conserved residues of the ABC signature motif, S142 and G144, into alanine. The G144A mutant is completely inactive and fails to dimerize, indicating an essential role of this residue in stabilizing the productive dimeric state. Mutant S142A retained considerable activity, and was able to dimerize, thus implying that the interaction of the serine with ATP is not essential for dimerization and catalysis. Furthermore, although the E166A and G144A mutants each alone are inactive, they produce an active heterodimer, showing that disruption of one active site can be tolerated. Our data suggest that ABC-ATPases with partially degenerated catalytic machineries, as they occur in vivo, can still form productive dimers to drive transport.

Adenosine Triphosphatases↗

PubMed2003

Archaeal homolog of bacterial type IV prepilin signal peptidases with broad substrate specificity.

A large number of secretory proteins in the thermoacidophile Sulfolobus solfataricus are synthesized as a precursor with an unusual leader peptide that resembles bacterial type IV prepilin signal sequences. This set of proteins includes the flagellin subunit but also various solute binding proteins. Here we describe the identification of the S. solfataricus homolog of bacterial type IV prepilin peptidases, termed PibD. PibD is an integral membrane protein that is phylogenetically related to the bacterial enzymes. When heterologously expressed in Escherichia coli, PibD is capable of processing both the flagellin and glucose-binding protein (GlcS) precursors. Site-directed mutagenesis of the GlcS signal peptide shows that the substrate specificity of PibD is consistent with the variations found in proteins with type IV prepilin-like signal sequences of S. solfataricus. We conclude that PibD is responsible for the processing of these secretory proteins in S. solfataricus.

Amino Acid Sequence↗

PubMed2002

Signal peptides of secreted proteins of the archaeon Sulfolobus solfataricus: a genomic survey.

Analysis of the recently completed genome sequence of the thermoacidophilic archaeon Sulfolobus solfataricus reveals that about 4.2% of its proteome consists of putative secretory proteins with signal peptides. This includes members of the four major classes of signal peptides: secretory signal peptides, twin-arginine signal peptides, possible lipoprotein precursors, and type IV pilin signal peptides. The latter group is surprisingly large compared to the size of the groups in other organisms and seems to be used predominately for a subset of extracellular substrate-binding proteins.

Amino Acid Sequence↗

PubMed2002

Sugar transport in (hyper)thermophilic archaea.

Hyperthermophilic archaea show important metabolic adaptations for growth on carbohydrates under hostile conditions. For carbohydrate uptake so far only ABC-type transporters have been described that are equipped with a uniquely high affinity as compared to mesophilic bacterial systems. This allows these organisms to efficiently scavenge all available carbohydrates from the extreme environment.

ATP-Binding Cassette Transporters↗

PubMed2002

The cell membrane plays a crucial role in survival of bacteria and archaea in extreme environments.

The cytoplasmic membrane of bacteria and archaea determine to a large extent the composition of the cytoplasm. Since the ion and in particular the proton and/or the sodium ion electrochemical gradients across the membranes are crucial for the bioenergetic conditions of these microorganisms, strategies are needed to restrict the permeation of these ions across their cytoplasmic membrane. The proton and sodium permeabilities of all biological membranes increase with the temperature. Psychrophilic and mesophilic bacteria, and mesophilic, (hyper)thermophilic and halophilic archaea are capable of adjusting the lipid composition of their membranes in such a way that the proton permeability at the respective growth temperature remains low and constant (homeo-proton permeability). Thermophilic bacteria, however, have more difficulties to restrict the proton permeation across their membrane at high temperatures and these organisms have to rely on the less permeable sodium ions for maintaining a high sodium-motive force for driving their energy requiring membrane-bound processes. Transport of solutes across the bacterial and archaeal membrane is mainly catalyzed by primary ATP driven transport systems or by proton or sodium motive force driven secondary transport systems. Unlike most bacteria, hyperthermophilic bacteria and archaea prefer primary ATP-driven uptake systems for their carbon and energy sources. Several high-affinity ABC transporters for sugars from hyperthermophiles have been identified and characterized. The activities of these ABC transporters allow these organisms to thrive in their nutrient-poor environments.

Archaea↗