Cloned (Comment) | Organism |
---|---|
recombinant expression of His-tagged enzyme with deletion of a membrane-anchoring region (residues 134-170) in Escherichia coli strain BL21(DE3) | Thermococcus onnurineus |
Crystallization (Comment) | Organism |
---|---|
purified isolated AAA+ module of LonB protease, mixing of 0.001 ml of 12 mg/ml protein solution in 10 mM Tris-HCl, pH 7.5, 100 mM NaCl, and 1 mM DTT, with 0.001 ml of reservoir solution containing 0.2 M potassium chloride, 0.01 M magnesium acetate, 0.05 M tri-sodium citrate dihydrate, pH 4.5, 12% PEG 4000, and 5% w/v n-dodecyl-beta-D-maltoside, 22°C, X-ray diffraction structure determination and analysis at 2.03 A resolution | Thermococcus onnurineus |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Thermococcus onnurineus | - |
- |
- |
Purification (Comment) | Organism |
---|---|
recombinant truncated His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity and anion exchange chromatography, the enzyme is reduced to the AAA+ domain by proteolytic treatment with trypsin followed by gel filtration | Thermococcus onnurineus |
Subunits | Comment | Organism |
---|---|---|
hexamer | - |
Thermococcus onnurineus |
Synonyms | Comment | Organism |
---|---|---|
lonB protease | - |
Thermococcus onnurineus |
TonLonB | - |
Thermococcus onnurineus |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
ATP | activates, the ATP-binding site is situated at the interface between the alpha/beta-subdomain and the alpha-subdomain. Motifs for ATP hydrolysis and nucleotide sensing are well conserved at the interface. Structure, detailed overview | Thermococcus onnurineus |
General Information | Comment | Organism |
---|---|---|
evolution | structural comparison of AAA+ modules between LonA and LonB reveals that the AAA+ modules of Lon proteases are separated into two distinct clades depending on their structural features. The H2-insert clade is characterized by the presence of both the PS-1 insertion and the H2 insertion. In the H2-insert clade, no member has an additional insertion, like Ins1, between alpha4 and beta2 in the RFC-B AAA+ module. Therefore, it is not appropriate to classify the AAA+ module of TonLonB as the H2-insert clade. Alternatively, the AAA+ module of LonB belongs to a new clade, named H2 & Ins1 insert clade is proposed. The AAA+ module of LonB belongs to the H2 & Ins1 insert clade (HINS clade), while the AAA+ module of LonA is a member of the HCLR clade | Thermococcus onnurineus |
additional information | crystal structure determination of AAA+ module. Structural basis for the ATP-independent proteolytic activity of LonB proteases and reclassification of their AAA+ modules, overview. The isolated AAA+ module, having no bound nucleotide, adopts a conformation virtually identical to the ADP-bound conformation of AAA+ modules in the hexameric structure of TonLonB. Despite the conservation of functional motifs, the iAAA+ module has no ATPase activity. In the hexameric conformation, an arginine finger (Arg311) in one AAA+ module stabilizes negative charge of gamma-phosphate of ATP bound to the adjacent AAA+ module, which is essential for ATP hydrolysis | Thermococcus onnurineus |