Single‐molecule analysis of dynamics and interactions of the SecYEG translocon

Koch, Sabrina; Seinen, Anne-Bart; Kamel, Michael; Kuckla, Daniel; Monzel, Cornelia; Kedrov, Alexej; Driessen, Arnold J. M.

doi:10.1111/febs.15596

Cited by 10 publications

(10 citation statements)

References 74 publications

(128 reference statements)

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“…The prominent effect of the lipid fatty acid composition on Sec-mediated translocation suggests that the hydrophobic core of the membrane could either affect stability and dynamics of the SecYEG translocon or be a novel factor that regulates SecA binding and SecA : SecYEG assembly. SecYEG is known to interact with specific lipids, such as PG and CL, and the interactions may cause heterogeneity in the structural dynamics [17][18][19]. To probe the effect of various environments on the translocon folding and stability, we established differential scanning fluorimetry (DSF) measurements, which report on the protein denaturation based on changes in the fluorescence emission of tryptophan residues [20].…”

Section: Secyeg Stability and Topology Are Not Affected By Fatty Acid...mentioning

confidence: 99%

Unsaturated fatty acids augment protein transport via the SecA:SecYEG translocon

et al. 2021

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The translocon SecYEG and the associated ATPase SecA form the primary protein secretion system in the cytoplasmic membrane of bacteria. The secretion is essentially dependent on the surrounding lipids, but the mechanistic understanding of their role in SecA : SecYEG activity is sparse. Here, we reveal that the unsaturated fatty acids (UFAs) of the membrane phospholipids, including tetraoleoyl-cardiolipin, stimulate SecA : SecYEGmediated protein translocation up to ten-fold. Biophysical analysis and molecular dynamics simulations show that UFAs increase the area per lipid and cause loose packing of lipid head groups, where the N-terminal amphipathic helix of SecA docks. While UFAs do not affect the translocon folding, they promote SecA binding to the membrane, and the effect is enhanced up to fivefold at elevated ionic strength. Tight SecA : lipid interactions convert into the augmented translocation. Our results identify the fatty acid structure as a notable factor in SecA : SecYEG activity, which may be crucial for protein secretion in bacteria, which actively change their membrane composition in response to their habitat.

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Section: Secyeg Stability and Topology Are Not Affected By Fatty Acid...mentioning

confidence: 99%

Unsaturated fatty acids augment protein transport via the SecA:SecYEG translocon

et al. 2021

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“…Several studies have applied total internal reflection fluorescent (TIRF) microscopy-based FRET measurement to characterize the coordination between channel opening, translocation, and ATP hydrolysis. [72][73][74][75] To better understand the conformational changes in SecY upon substrate engagement, Collinson, Tuma and colleagues investigated SecY pore dynamics using TIRF-based FRET measurements and the model substrate proOmpA (Figure 5A). [74] SecY and SecA were labeled with the FRET pair Cy3 and Cy5, respectively, in positions that report on whether the channel is plugged (Figure 5B).…”

Section: Single-molecule Fretmentioning

confidence: 99%

“…While the unlocked state had been predicted by purely structural characterization, [76] the single molecule experiment allowed direct observation of this transition, its dynamics, and its dependence on ATP. [75] Rapoport and colleagues also used single-molecule FRET to monitor the role of SecA ATPase activity during translocation. [73] SecY was donor-labeled at the periplasmic end of the channel.…”

Section: Single-molecule Fretmentioning

confidence: 99%

Methodologies for Measuring Protein Trafficking across Cellular Membranes

Lyu

Genereux

2021

ChemPlusChem

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Nearly all proteins are synthesized in the cytosol. The majority of this proteome must be trafficked elsewhere, such as to membranes, to subcellular compartments, or outside of the cell. Proper trafficking of nascent protein is necessary for protein folding, maturation, quality control and cellular and organismal health. To better understand cellular biology, molecular and chemical technologies to properly characterize protein trafficking (and mistrafficking) have been developed and applied. Herein, we take a biochemical perspective to review technologies that enable spatial and temporal measurement of protein distribution, focusing on both the most widely adopted methodologies and exciting emerging approaches.

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“…The cell has in place membrane proteins which form channels that allow passive or active diffusion of molecules. The inserted channels can result from the multimerization/oligomerisation of a single protein such as TatA [2], although it is likely that parts of TatB and TatC also form the channel, or the hetero-oligomerisation of several subunits such as the translocon SecYEG [3], both pores being involved in protein translocation. Alternatively, a single protein that has several transmembrane segments can fold in the membrane so that it builds a channel structure, such as LacY [4], a permease that allows the passage of lactose.…”

Section: Introductionmentioning

confidence: 99%

Bacterial protein secretion systems: Game of types

Filloux

2022

Microbiology

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Protein trafficking across the bacterial envelope is a process that contributes to the organisation and integrity of the cell. It is the foundation for establishing contact and exchange between the environment and the cytosol. It helps cells to communicate with one another, whether they establish symbiotic or competitive behaviours. It is instrumental for pathogenesis and for bacteria to subvert the host immune response. Understanding the formation of envelope conduits and the manifold strategies employed for moving macromolecules across these channels is a fascinating playground. The diversity of the nanomachines involved in this process logically resulted in an attempt to classify them, which is where the protein secretion system types emerged. As our knowledge grew, so did the number of types, and their rightful nomenclature started to be questioned. While this may seem a semantic or philosophical issue, it also reflects scientific rigour when it comes to assimilating findings into textbooks and science history. Here I give an overview on bacterial protein secretion systems, their history, their nomenclature and why it can be misleading for newcomers in the field. Note that I do not try to suggest a new nomenclature. Instead, I explore the reasons why naming could have escaped our control and I try to reiterate basic concepts that underlie protein trafficking cross membranes.

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Single‐molecule analysis of dynamics and interactions of the SecYEG translocon

Cited by 10 publications

References 74 publications

Unsaturated fatty acids augment protein transport via the SecA:SecYEG translocon

Unsaturated fatty acids augment protein transport via the SecA:SecYEG translocon

Methodologies for Measuring Protein Trafficking across Cellular Membranes

Bacterial protein secretion systems: Game of types

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