Cloned (Comment) | Organism |
---|---|
expression of His6-tagged wild-type and mutant EndAs in Escherichi coli strain Rosetta 2(DE3) | Aeropyrum pernix |
Crystallization (Comment) | Organism |
---|---|
purified recombinant wild-type and mutant EndAs, method screening, sitting drop vapor diffusion technique, mixing of 0.001 ml protein solution with 0.001 ml reservoir solution containing 0.2 M NaCl, 0.1 M phosphate-citrate, pH 4.2, and 10% w/v PEG 3000, equilibration over 0.1 ml reservoir solution, 22°C, X-ray diffraction structure determmination and analysis at 1.7-2.3 A resolution, molecular replacement | Aeropyrum pernix |
Protein Variants | Comment | Organism |
---|---|---|
D49A | site-directed mutagenesis, the mutation does not affect the enzyme activity | Aeropyrum pernix |
E43A | site-directed mutagenesis, the mutation does not affect the enzyme activity | Aeropyrum pernix |
E51A | site-directed mutagenesis, the mutation does not affect the enzyme activity | Aeropyrum pernix |
F50A | site-directed mutagenesis, the mutant shows reduced enzyme activity compared to the wild-type enzyme | Aeropyrum pernix |
H133A | site-directed mutagenesis, a catalytic site mutant, crystal structure determination | Aeropyrum pernix |
K44A | site-directed mutagenesis, the mutant shows almost no enzyme activity | Aeropyrum pernix |
P45A | site-directed mutagenesis, the mutant shows reduced enzyme activity compared to the wild-type enzyme | Aeropyrum pernix |
R46A | site-directed mutagenesis, the mutant shows reduced enzyme activity compared to the wild-type enzyme | Aeropyrum pernix |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Aeropyrum pernix | Q9YBF1 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and mutant EndAs from Escherichia coli strain Rosetta 2(DE3) by heat treatment at 70°C for 30 min, followed by metal affinity chromatography and gel filtration | Aeropyrum pernix |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
additional information | Sulfolobus tokodaii tRNATrp precursor as a substrate | Aeropyrum pernix | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
EndA | - |
Aeropyrum pernix |
splicing endonuclease | - |
Aeropyrum pernix |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
70 | - |
assay at | Aeropyrum pernix |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Aeropyrum pernix |
General Information | Comment | Organism |
---|---|---|
evolution | the crenarchaeal heterotetrameric EndAs can be further classified into two subfamilies based on the size of the structural subunit. Subfamily A possesses a structural subunit similar in size to the catalytic subunit, whereas subfamily B possesses a structural subunit significantly smaller than the catalytic subunit | Aeropyrum pernix |
additional information | EndA from Aeropyrum pernix also possesses an extra loop region that is characteristic of crenarchaeal EndAs, the conserved lysine residue Lys44 in the loop is important for endonuclease activity, substrate docking modeling, overview | Aeropyrum pernix |