Julia Oswald

Bio: Julia Oswald is an academic researcher from University of Freiburg. The author has contributed to research in topics: Sec61 & Membrane protein. The author has an hindex of 2, co-authored 2 publications receiving 11 citations.

The Dynamic SecYEG Translocon.

Abstract: The spatial and temporal coordination of protein transport is an essential cornerstone of the bacterial adaptation to different environmental conditions. By adjusting the protein composition of extra-cytosolic compartments, like the inner and outer membranes or the periplasmic space, protein transport mechanisms help shaping protein homeostasis in response to various metabolic cues. The universally conserved SecYEG translocon acts at the center of bacterial protein transport and mediates the translocation of newly synthesized proteins into and across the cytoplasmic membrane. The ability of the SecYEG translocon to transport an enormous variety of different substrates is in part determined by its ability to interact with multiple targeting factors, chaperones and accessory proteins. These interactions are crucial for the assisted passage of newly synthesized proteins from the cytosol into the different bacterial compartments. In this review, we summarize the current knowledge about SecYEG-mediated protein transport, primarily in the model organism Escherichia coli, and describe the dynamic interaction of the SecYEG translocon with its multiple partner proteins. We furthermore highlight how protein transport is regulated and explore recent developments in using the SecYEG translocon as an antimicrobial target.

Eeyarestatin 24 Impairs SecYEG-dependent Protein Trafficking and Inhibits Growth of Clinically Relevant Pathogens

Abstract: Eeyarestatin 1 (ES1) is an inhibitor of endoplasmic reticulum (ER) associated protein degradation, Sec61-dependent Ca2+ homeostasis and protein translocation into the ER. Recently, evidence was presented showing that a smaller analog of ES1, ES24, targets the Sec61-translocon, and captures it in an open conformation that is translocation-incompetent. We now show that ES24 impairs protein secretion and membrane protein insertion in Escherichia coli via the homologous SecYEG-translocon. Transcriptomic analysis suggested that ES24 has a complex mode of action, probably involving multiple targets. Interestingly, ES24 shows antibacterial activity toward clinically relevant strains. Furthermore, the antibacterial activity of ES24 is equivalent to or better than that of nitrofurantoin, a known antibiotic that, although structurally similar to ES24, does not interfere with SecYEG-dependent protein trafficking. Like nitrofurantoin, we find that ES24 requires activation by the NfsA and NfsB nitroreductases, suggesting that the formation of highly reactive nitroso intermediates is essential for target inactivation in vivo.

The Dynamic SecYEG Translocon.

Abstract: The spatial and temporal coordination of protein transport is an essential cornerstone of the bacterial adaptation to different environmental conditions. By adjusting the protein composition of extra-cytosolic compartments, like the inner and outer membranes or the periplasmic space, protein transport mechanisms help shaping protein homeostasis in response to various metabolic cues. The universally conserved SecYEG translocon acts at the center of bacterial protein transport and mediates the translocation of newly synthesized proteins into and across the cytoplasmic membrane. The ability of the SecYEG translocon to transport an enormous variety of different substrates is in part determined by its ability to interact with multiple targeting factors, chaperones and accessory proteins. These interactions are crucial for the assisted passage of newly synthesized proteins from the cytosol into the different bacterial compartments. In this review, we summarize the current knowledge about SecYEG-mediated protein transport, primarily in the model organism Escherichia coli, and describe the dynamic interaction of the SecYEG translocon with its multiple partner proteins. We furthermore highlight how protein transport is regulated and explore recent developments in using the SecYEG translocon as an antimicrobial target.

Membrane protein biogenesis at the ER: the highways and byways.

Abstract: The Sec61 complex is the major protein translocation channel of the endoplasmic reticulum (ER), where it plays a central role in the biogenesis of membrane and secretory proteins. Whilst Sec61-mediated protein translocation is typically coupled to polypeptide synthesis, suggestive of significant complexity, an obvious characteristic of this core translocation machinery is its surprising simplicity. Over thirty years after its initial discovery, we now understand that the Sec61 complex is in fact the central piece of an elaborate jigsaw puzzle, which can be partly solved using new research findings. We propose that the Sec61 complex acts as a dynamic hub for co-translational protein translocation at the ER, proactively recruiting a range of accessory complexes that enhance and regulate its function in response to different protein clients. It is now clear that the Sec61 complex does not have a monopoly on co-translational insertion, with some transmembrane proteins preferentially utilising the ER membrane complex instead. We also have a better understanding of post-insertion events, where at least one membrane-embedded chaperone complex can capture the newly inserted transmembrane domains of multi-span proteins and co-ordinate their assembly into a native structure. Having discovered this array of Sec61-associated components and competitors, our next challenge is to understand how they act together in order to expand the range and complexity of the membrane proteins that can be synthesised at the ER. Furthermore, this diversity of components and pathways may open up new opportunities for targeted therapeutic interventions designed to selectively modulate protein biogenesis at the ER.

Inhibitors of the Sec61 Complex and Novel High Throughput Screening Strategies to Target the Protein Translocation Pathway.

Abstract: Proteins targeted to the secretory pathway start their intracellular journey by being transported across biological membranes such as the endoplasmic reticulum (ER). A central component in this protein translocation process across the ER is the Sec61 translocon complex, which is only intracellularly expressed and does not have any enzymatic activity. In addition, Sec61 translocon complexes are difficult to purify and to reconstitute. Screening for small molecule inhibitors impairing its function has thus been notoriously difficult. However, such translocation inhibitors may not only be valuable tools for cell biology, but may also represent novel anticancer drugs, given that cancer cells heavily depend on efficient protein translocation into the ER to support their fast growth. In this review, different inhibitors of protein translocation will be discussed, and their specific mode of action will be compared. In addition, recently published screening strategies for small molecule inhibitors targeting the whole SRP-Sec61 targeting/translocation pathway will be summarized. Of note, slightly modified assays may be used in the future to screen for substances affecting SecYEG, the bacterial ortholog of the Sec61 complex, in order to identify novel antibiotic drugs.

Stress-Based High-Throughput Screening Assays to Identify Inhibitors of Cell Envelope Biogenesis.

Abstract: The structural integrity of the Gram-negative cell envelope is guarded by several stress responses, such as the σE, Cpx and Rcs systems. Here, we report on assays that monitor these responses in E. coli upon addition of antibacterial compounds. Interestingly, compromised peptidoglycan synthesis, outer membrane biogenesis and LPS integrity predominantly activated the Rcs response, which we developed into a robust HTS (high-throughput screening) assay that is suited for phenotypic compound screening. Furthermore, by interrogating all three cell envelope stress reporters, and a reporter for the cytosolic heat-shock response as control, we found that inhibitors of specific envelope targets induce stress reporter profiles that are distinct in quality, amplitude and kinetics. Finally, we show that by using a host strain with a more permeable outer membrane, large-scaffold antibiotics can also be identified by the reporter assays. Together, the data suggest that stress profiling is a useful first filter for HTS aimed at inhibitors of cell envelope processes.

How do Chaperones Bind (Partly) Unfolded Client Proteins

Abstract: Molecular chaperones are central to cellular protein homeostasis. Dynamic disorder is a key feature of the complexes of molecular chaperones and their client proteins, and it facilitates the client release towards a folded state or the handover to downstream components. The dynamic nature also implies that a given chaperone can interact with many different client proteins, based on physico-chemical sequence properties rather than on structural complementarity of their (folded) 3D structure. Yet, the balance between this promiscuity and some degree of client specificity is poorly understood. Here, we review recent atomic-level descriptions of chaperones with client proteins, including chaperones in complex with intrinsically disordered proteins, with membrane-protein precursors, or partially folded client proteins. We focus hereby on chaperone-client interactions that are independent of ATP. The picture emerging from these studies highlights the importance of dynamics in these complexes, whereby several interaction types, not only hydrophobic ones, contribute to the complex formation. We discuss these features of chaperone-client complexes and possible factors that may contribute to this balance of promiscuity and specificity.