scholarly journals Homologous bd oxidases share the same architecture but differ in mechanism

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Alexander Theßeling ◽  
Tim Rasmussen ◽  
Sabrina Burschel ◽  
Daniel Wohlwend ◽  
Jan Kägi ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Specific Inhibitor ◽  
Protonmotive Force ◽  
Geobacillus Thermodenitrificans ◽  
E Coli ◽  
Cryo Electron Microscopy ◽  
Respiratory Chains

Abstract Cytochrome bd oxidases are terminal reductases of bacterial and archaeal respiratory chains. The enzyme couples the oxidation of ubiquinol or menaquinol with the reduction of dioxygen to water, thus contributing to the generation of the protonmotive force. Here, we determine the structure of the Escherichia coli bd oxidase treated with the specific inhibitor aurachin by cryo-electron microscopy (cryo-EM). The major subunits CydA and CydB are related by a pseudo two fold symmetry. The heme b and d cofactors are found in CydA, while ubiquinone-8 is bound at the homologous positions in CydB to stabilize its structure. The architecture of the E. coli enzyme is highly similar to that of Geobacillus thermodenitrificans, however, the positions of heme b595 and d are interchanged, and a common oxygen channel is blocked by a fourth subunit and substituted by a more narrow, alternative channel. Thus, with the same overall fold, the homologous enzymes exhibit a different mechanism.

scholarly journals Assembly principles of a unique cage formed by hexameric and decameric E. coli proteins

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Hélène Malet ◽  
Kaiyin Liu ◽  
Majida El Bakkouri ◽  
Sze Wah Samuel Chan ◽  
Gregory Effantin ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Lysine Decarboxylase ◽  
Lateral Interactions ◽  
E Coli ◽  
Aaa Atpase ◽  
Cryo Electron Microscopy ◽  
Conformational Rearrangements ◽  
The Individual ◽  
Assembly Principles

A 3.3 MDa macromolecular cage between two Escherichia coli proteins with seemingly incompatible symmetries–the hexameric AAA+ ATPase RavA and the decameric inducible lysine decarboxylase LdcI–is reconstructed by cryo-electron microscopy to 11 Å resolution. Combined with a 7.5 Å resolution reconstruction of the minimal complex between LdcI and the LdcI-binding domain of RavA, and the previously solved crystal structures of the individual components, this work enables to build a reliable pseudoatomic model of this unusual architecture and to identify conformational rearrangements and specific elements essential for complex formation. The design of the cage created via lateral interactions between five RavA rings is unique for the diverse AAA+ ATPase superfamily.

scholarly journals Cardiolipin enhances the enzymatic activity of cytochrome bd and cytochrome bo3 solubilized in dodecyl-maltoside

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Amer H. Asseri ◽  
Albert Godoy-Hernandez ◽  
Hojjat Ghasemi Goojani ◽  
Holger Lill ◽  
Junshi Sakamoto ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Oxygen Consumption ◽  
Integral Membrane Proteins ◽  
Terminal Oxidase ◽  
Geobacillus Thermodenitrificans ◽  
E Coli ◽  
Cytochrome Bd ◽  
Cytochrome Bo3 ◽  
Dodecyl Maltoside ◽  
Consumption Activity

AbstractCardiolipin (CL) is a lipid that is found in the membranes of bacteria and the inner membranes of mitochondria. CL can increase the activity of integral membrane proteins, in particular components of respiratory pathways. We here report that CL activated detergent-solubilized cytochrome bd, a terminal oxidase from Escherichia coli. CL enhanced the oxygen consumption activity ~ twofold and decreased the apparent KM value for ubiquinol-1 as substrate from 95 µM to 35 µM. Activation by CL was also observed for cytochrome bd from two Gram-positive species, Geobacillus thermodenitrificans and Corynebacterium glutamicum, and for cytochrome bo3 from E. coli. Taken together, CL can enhance the activity of detergent-solubilized cytochrome bd and cytochrome bo3.

Escherichia coli 70 S ribosome at 15 Å resolution by cryo-electron microscopy: localization of fmet-tRNAfMet and fitting of L1 protein

1998 ◽  
Vol 280 (1) ◽  
pp. 103-116 ◽  
Author(s):  
Arun Malhotra ◽  
Pawel Penczek ◽  
Rajendra K Agrawal ◽  
Irene S Gabashvili ◽  
Robert A Grassucci ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Cryo Electron Microscopy ◽  
L1 Protein

scholarly journals In Vivo Modulation of a DnaJ Homolog, CbpA, by CbpM

2007 ◽  
Vol 189 (9) ◽  
pp. 3635-3638 ◽  
Author(s):  
Matthew R. Chenoweth ◽  
Nancy Trun ◽  
Sue Wickner
Keyword(s):  
Escherichia Coli ◽  
Cell Growth ◽  
Stationary Phase ◽  
Specific Inhibitor ◽  
Vitro Activity ◽  
In Vitro Activity ◽  
E Coli ◽  
Hsp70 Chaperone

ABSTRACT CbpA, an Escherichia coli DnaJ homolog, can function as a cochaperone for the DnaK/Hsp70 chaperone system, and its in vitro activity can be modulated by CbpM. We discovered that CbpM specifically inhibits the in vivo activity of CbpA, preventing it from functioning in cell growth and division. Furthermore, we have shown that CbpM interacts with CbpA in vivo during stationary phase, suggesting that the inhibition of activity is a result of the interaction. These results reveal that the activity of the E. coli DnaK system can be regulated in vivo by a specific inhibitor.

In Vitro Antibacterial Activity and Mechanism of Monocaprylin against Escherichia coli and Staphylococcus aureus

2018 ◽  
Vol 81 (12) ◽  
pp. 1988-1996 ◽  
Author(s):  
JIANYU WANG ◽  
MAOMAO MA ◽  
JUN YANG ◽  
LONG CHEN ◽  
PING YU ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antibacterial Activity ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Sodium Benzoate ◽  
Rapid Change ◽  
Potassium Sorbate ◽  
E Coli

ABSTRACT In the present study, the antibacterial activity of monocaprylin in comparison with sodium benzoate and potassium sorbate against Staphylococcus aureus and Escherichia coli was assessed by measuring MIC, MBC, effect of pH on MIC, and incubation temperature on bactericidal efficacy. Results showed that monocaprylin exhibited an excellent antibacterial activity against both strains, with the lowest MIC and MBC of 1.28 mg/mL. A MIC of monocaprylin remained unchanged despite the pH values of culture medium, ranging from 5 to 9, unlike that of potassium sorbate or sodium benzoate. Furthermore, monocaprylin at MBC effectively reduced the population of E. coli and S. aureus by >5.5 log CFU/mL at 25°C within 6 h and decreased E. coli by approximately 5.0 log CFU/mL and S. aureus by 2.9 log CFU/mL at 12 h. The underlying mechanism of monocaprylin was then investigated by measuring β-galactosidase activity, membrane potential, release of cellular contents, scanning electron microscopy, and transmission electron microscopy observations. Results indicated that monocaprylin killed E. coli by the rapid change in permeability and integrity of cell membrane, leading to decline of membrane potential, leakage of nucleic acids and proteins, and ultimately cell membrane disintegration and lysis. On the other hand, monocaprylin might exert its antibacterial activity against S. aureus mainly by diffusing across the cell wall, collapsing the cell membrane, and disturbing the order of intracellular contents. These findings indicated that monocaprylin had better antibacterial ability compared with traditional synthetic preservatives and might be a potential antibacterial additive independent of pH.

scholarly journals Slow Polymerization of Mycobacterium tuberculosis FtsZ

2000 ◽  
Vol 182 (14) ◽  
pp. 4028-4034 ◽  
Author(s):  
E. Lucile White ◽  
Larry J. Ross ◽  
Robert C. Reynolds ◽  
Lainne E. Seitz ◽  
Georgia D. Moore ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Mycobacterium Tuberculosis ◽  
Tubulin Polymerization ◽  
Gtpase Activity ◽  
Gtp Hydrolysis ◽  
Minimum Concentration ◽  
E Coli ◽  
Cell Division Protein ◽  
Mycobacterial Protein

ABSTRACT The essential cell division protein, FtsZ, from Mycobacterium tuberculosis has been expressed in Escherichia coliand purified. The recombinant protein has GTPase activity typical of tubulin and other FtsZs. FtsZ polymerization was studied using 90° light scattering. The mycobacterial protein reaches maximum polymerization much more slowly (∼10 min) than E. coliFtsZ. Depolymerization also occurs slowly, taking 1 h or longer under most conditions. Polymerization requires both Mg2+and GTP. The minimum concentration of FtsZ needed for polymerization is 3 μM. Electron microscopy shows that polymerized M. tuberculosis FtsZ consists of strands that associate to form ordered aggregates of parallel protofilaments. Ethyl 6-amino-2,3-dihydro-4-phenyl-1H-pyrido[4,3-b][1,4]diazepin-8-ylcarbamate (SRI 7614), an inhibitor of tubulin polymerization synthesized at Southern Research Institute, inhibits M. tuberculosis FtsZ polymerization, inhibits GTP hydrolysis, and reduces the number and sizes of FtsZ polymers.

scholarly journals The Cryo-Electron Microscopy Structure of the Type 1 Chaperone-Usher Pilus Rod

2017 ◽  
Author(s):  
Manuela K. Hospenthal ◽  
Tiago R. D. Costa ◽  
Adam Redzej ◽  
James Lillington ◽  
Gabriel Waksman
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
Urinary Tract ◽  
Virulence Factors ◽  
Biomechanical Properties ◽  
Bacterial Attachment ◽  
Host Cells ◽  
Hostile Environment ◽  
Cryo Electron Microscopy

ABSTRACTChaperone-usher pili are long, polymeric protein fibres displayed on the surface of many bacterial pathogens. These critical virulence factors allow bacteria to specifically attach to host cells during infection. The type 1 and P pili of uropathogenicEscherichia coli(UPEC) play important roles during UPEC’s colonisation of the urinary tract, mediating bacterial attachment to the bladder and kidney, respectively. Also, their biomechanical properties that allow them to reversibly uncoil in response to flow-induced forces are critical for UPEC’s ability to retain a foothold in the unique and hostile environment of the urinary tract. Here we provide the 4.2 Å resolution cryo-electron microscopy (cryo-EM) structure of the type 1 pilus rod, which together with the previous structure of the P pilus rod, enables us to understand the remarkable “spring-like” properties of chaperone-usher pili in more detail.

scholarly journals High-resolution cryo-electron microscopy structure of the Escherichia coli 50S subunit and validation of nucleotide modifications

2019 ◽  
Author(s):  
Vanja Stojković ◽  
Alexander G. Myasnikov ◽  
Iris D. Young ◽  
Adam Frost ◽  
James S. Fraser ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Escherichia Coli ◽  
High Resolution ◽  
Rna Structure ◽  
Ribosome Biogenesis ◽  
Precise Determination ◽  
23S Rrna ◽  
Modified Nucleotides ◽  
X Ray Crystallography ◽  
Cryo Electron Microscopy

ABSTRACTPost-transcriptional ribosomal RNA (rRNA) modifications are present in all organisms, but their exact functional roles and positions are yet to be fully characterized. Modified nucleotides have been implicated in the stabilization of RNA structure and regulation of ribosome biogenesis and protein synthesis. In some instances, rRNA modifications can confer antibiotic resistance. High-resolution ribosome structures are thus necessary for precise determination of modified nucleotides’ positions, a task that has previously been accomplished by X-ray crystallography. Here we present a cryo-electron microscopy (cryo-EM) structure of Escherichia coli (E. coli) 50S subunit at an average resolution of 2.2Å as an additional approach for mapping modification sites. Our structure confirms known modifications present in 23S rRNA and additionally allows for localization of Mg2+ ions and their coordinated water molecules. Using our cryo-EM structure as a testbed, we developed a program for identification of post-transcriptional rRNA modifications using a cryo-EM map. This program can be easily used on any RNA-containing cryo-EM structure, and an associated Coot plugin allows for visualization of validated modifications, making it highly accessible.

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