Abstract:Here we report the discovery of a bacterial DNA-segregating actinlike protein (BtParM) from Bacillus thuringiensis, which forms novel antiparallel, two-stranded, supercoiled, nonpolar helical filaments, as determined by electron microscopy. The BtParM filament features of supercoiling and forming antiparallel double-strands are unique within the actin fold superfamily, and entirely different to the straight, double-stranded, polar helical filaments of all other known ParMs and of eukaryotic F-actin. The BtParM… Show more
“…While a few actin‐like filaments have been reported to be single stranded (crenactin) or four‐stranded (Alp12, BtParM), most of them possess a staggered double‐stranded arrangement, with intimate contacts between strands. In contrast, the MamK filament is nonstaggered, with a large gap between adjacent strands [Fig.…”
Magnetotactic bacteria possess cellular compartments called magnetosomes that sense magnetic fields. Alignment of magnetosomes in the bacterial cell is necessary for their function, and this is achieved through anchoring of magnetosomes to filaments composed of the protein MamK. MamK is an actin homolog that polymerizes upon ATP binding. Here, we report the structure of the MamK filament at~6.5 Å , obtained by cryo-Electron Microscopy. This structure confirms our previously reported double-stranded, nonstaggered architecture, and reveals the molecular basis for filament formation. While MamK is closest in sequence to the bacterial actin MreB, the longitudinal contacts along each MamK strand most closely resemble those of eukaryotic actin. In contrast, the cross-strand interface, with a surprisingly limited set of contacts, is novel among actin homologs and gives rise to the nonstaggered architecture.
“…While a few actin‐like filaments have been reported to be single stranded (crenactin) or four‐stranded (Alp12, BtParM), most of them possess a staggered double‐stranded arrangement, with intimate contacts between strands. In contrast, the MamK filament is nonstaggered, with a large gap between adjacent strands [Fig.…”
Magnetotactic bacteria possess cellular compartments called magnetosomes that sense magnetic fields. Alignment of magnetosomes in the bacterial cell is necessary for their function, and this is achieved through anchoring of magnetosomes to filaments composed of the protein MamK. MamK is an actin homolog that polymerizes upon ATP binding. Here, we report the structure of the MamK filament at~6.5 Å , obtained by cryo-Electron Microscopy. This structure confirms our previously reported double-stranded, nonstaggered architecture, and reveals the molecular basis for filament formation. While MamK is closest in sequence to the bacterial actin MreB, the longitudinal contacts along each MamK strand most closely resemble those of eukaryotic actin. In contrast, the cross-strand interface, with a surprisingly limited set of contacts, is novel among actin homologs and gives rise to the nonstaggered architecture.
“…These designs are likely to represent just a fraction of the total filament architectures formed from the actin fold. Negative stain EM data point to other designs that are comprised of four filaments arranged as closed or open tubules, thus we expect the wider application of high resolution cryoEM to uncover many more architectures.…”
Structural biology has experienced several transformative technological advances in recent years. These include: development of extremely bright Xray sources (microfocus synchrotron beamlines and free electron lasers) and the use of electrons to extend protein crystallography to ever decreasing crystal sizes; and an increase in the resolution attainable by cryo-electron microscopy. Here we discuss the use of these techniques in general terms and highlight their application for biological filament systems, an area that is severely underrepresented in atomic resolution structures. We assemble a model of a capped tropomyosin-actin minifilament to demonstrate the utility of combining structures determined by different techniques. Finally, we survey the methods that attempt to transform high resolution structural biology into more physiological environments, such as the cell. Together these techniques promise a compelling decade for structural biology and, more importantly, they will provide exciting discoveries in understanding the designs and purposes of biological machines.
“…Figure 1.Two views of the structures of filaments formed from the actin (blue/orange) and tubulin (red/yellow) folds. The actins are: the twisted single-stranded crenactin from the archaeon Pyrobaculum calidifontis , 33,39
Caulobacter crescentus MreB filament formed from an antiparallel non-twisted pair of strands ( Cc MreB), 23 right-handed eukaryotic F-actin, 10 left-handed Escherichia coli ParM from the R1 plasmid ( Ec ParM), 35
Clostridium tetani open nanotubules from the pE88 plasmid ( Ct ParM) which are 2 antiparallel related copies of a parallel pair of strands 31 and Bacillus thuringiensis supercoiled antiparallel filaments and nanotubules from the pBMB67 plasmid ( Bt ParM) 34 . The tubulins are: the eukaryotic microtubule, 40
Bacillus thuringiensis TubZ from the pBtoxis plasmid ( Bt TubZ) 41 and Methanococcus jannaschii FtsZ ( Mj FtsZ) 42 .…”
Section: Variety In Prokaryotic Actin-like Filamentsmentioning
confidence: 99%
“…1), a far departure from the construction of F-actin or any other ParM 34 . ParM's are plasmid segregation polymerizing actins that are linked to the parC regions of the plasmid DNA via adaptor proteins (ParRs), which are specific to each individual ParM/ parC combination.…”
Section: Variety In Prokaryotic Actin-like Filamentsmentioning
confidence: 99%
“…2). 34 Thus in the case of Bt ParCMR, the unique Bt ParM nanotubule geometry requires a second component of the Bt ParCMR system, ParR or ParR/ parC . Figure 2.In the presence of molecular crowding Bt ParM formed rafts of filaments lying in parallel as observed by electron microscopy (A).…”
Section: Variety In Prokaryotic Actin-like Filamentsmentioning
From yeast to man, an evolutionary distance of 1.3 billion years, the F-actin filament structure has been conserved largely in line with the 94% sequence identity. The situation is entirely different in bacteria. In comparison to eukaryotic actins, the bacterial actin-like proteins (ALPs) show medium to low levels of sequence identity. This is extreme in the case of the ParM family of proteins, which often display less than 20% identity. ParMs are plasmid segregation proteins that form the polymerizing motors that propel pairs of plasmids to the extremities of a cell prior to cell division, ensuring faithful inheritance of the plasmid. Recently, exotic ParM filament structures have been elucidated that show ParM filament geometries are not limited to the standard polar pair of strands typified by actin. Four-stranded non-polar ParM filaments existing as open or closed nanotubules are found in Clostridium tetani and Bacillus thuringiensis, respectively. These diverse architectures indicate that the actin fold is capable of forming a large variety of filament morphologies, and that the conception of the “actin” filament has been heavily influenced by its conservation in eukaryotes. Here, we review the history of the structure determination of the eukaryotic actin filament to give a sense of context for the discovery of the new ParM filament structures. We describe the novel ParM geometries and predict that even more complex actin-like filaments may exist in bacteria. Finally, we compare the architectures of filaments arising from the actin and tubulin folds and conclude that the basic units possess similar properties that can each form a range of structures. Thus, the use of the actin fold in microfilaments and the tubulin fold for microtubules likely arose from a wider range of filament possibilities, but became entrenched as those architectures in early eukaryotes.
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