Activating Compound | Comment | Organism | Structure |
---|---|---|---|
cardiolipin | stimulates activity of full-length GspEEpsE when in complex with the cytoplasmic domain of GspLEpsL | Vibrio cholerae | |
additional information | hexamerization enhances the ATPase activity of recombinant DN1GspEEpsE-linker-Hcp1 variants compared to monomeric DN1GspEEpsE by more than 20fold | Vibrio cholerae |
Cloned (Comment) | Organism |
---|---|
recombinant expression of the enzyme GspE as fusion proteins with Pseudomonas aeruginosa Hcp1, i.e. DELTAN1GspEEpsE-KLASGHcp1, DELTAN1GspEEpsE-GSGSGS-Hcp1, DELTAN1GspEEpsE-KLASGAGHcp1, and DELTAN1GspEEpsE-KLASGAGH-Hcp1 showing homogeneous hexamer formation, several variants of DN1GspEEpsE-Hcp1 fusions with different linker sequences are constructed | Vibrio cholerae |
Crystallization (Comment) | Organism |
---|---|
recombinant enzyme GspE-Hcp1 fusion protein hexamers, sitting drop vapour diffusion method, mixing of 0.001 ml protein solution containing 20 mM Tris-HCl, pH 8.0, 500 mM NaCl, 5% v/v glycerol, 1 mM TCEP/HCl, 5 mM AMPPNP, and 5 mM MgCl2 with different precipitant solutions, containing for DN1GspEEpsE-GSGSGS-Hcp1 7% PEG 6000 and 0.1 M bicine, pH 9.0, for DN1GspEEpsE-KLASGAGH-Hcp1 16% PEG 300, 0.2 M ammonium sulfate, 0.1 M Bis Tris, pH 6.1, 5 mM ADP, 5 mM MgCl2, 5 mM AlCl3, and 15 mM NaF, for DELTAN1GspEEpsE-KLASGAG-Hcp1 7% PEG 3350, 0.12 M ammonium citrate pH 7.0, 5 mM ADP, and 5 mM MgCl2, and for DELTAN1GspEEpsE-KLASG-Hcp1 12.5% PEG 20000, 0.1 M bicine pH 9.0, and 2% v/v 1,4-dioxane, at 4°C, X-ray diffraction structure determination and analysis at 4.09-7.6 A resolution | Vibrio cholerae |
Protein Variants | Comment | Organism |
---|---|---|
additional information | four hexamers of Vibrio cholerae GspEEpsE are obtained when fused to Hcp1 as an assistant hexamer, shown with native mass spectrometry. The enzymatic activity of the GspEEpsE-Hcp1 fusions is about 20 times higher than that of a GspEEpsE monomer. Crystal structures of GspEEpsE-Hcp1 fusions with different linker lengths reveal regular and elongated hexamers of GspEEpsE with major differences in domain orientation within subunits, and in subunit assembly | Vibrio cholerae |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | required | Vibrio cholerae |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
56000 | - |
x * 56000 | Vibrio cholerae |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + H2O | Vibrio cholerae | - |
ADP + phosphate | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Vibrio cholerae | P37093 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
ATP + H2O | - |
Vibrio cholerae | ADP + phosphate | - |
? |
Subunits | Comment | Organism |
---|---|---|
? | x * 56000 | Vibrio cholerae |
More | recombinant enzyme GspE-Hcp1 fusion proteins, i.e. DELTAN1GspEEpsE-KLASGHcp1, DELTAN1GspEEpsE-GSGSGS-Hcp1, DELTAN1GspEEpsE-KLASGAGHcp1, and DELTAN1GspEEpsE-KLASGAGH-Hcp1, show homogeneous hexamer formation, structure analysis, detailed overview. The hexamerization enhances the ATPase activity of these four DN1GspEEpsE-linker-Hcp1 variants compared to monomeric DN1GspEEpsE by more than 20fold. The metal binding domains are located on the periphery of both DN1GspEEpsE hexamers | Vibrio cholerae |
Synonyms | Comment | Organism |
---|---|---|
critical multidomain T2SS assembly ATPase | - |
Vibrio cholerae |
GspE | - |
Vibrio cholerae |
GspEEpsE | - |
Vibrio cholerae |
type II secretion ATPase | - |
Vibrio cholerae |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Vibrio cholerae |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8.5 | - |
assay at | Vibrio cholerae |
General Information | Comment | Organism |
---|---|---|
evolution | the T2SS secretion ATPase GspE belongs to the family of Type II/IV secretion ATPases | Vibrio cholerae |
metabolism | the type II secretion system (T2SS), a multiprotein machinery spanning two membranes in Gram-negative bacteria, incudes the critical multidomain GspEEpsE, and is responsible for the secretion of folded proteins from the periplasm across the outer membrane | Vibrio cholerae |