Literature summary for 4.6.1.16 extracted from

Okuda, M.; Shiba, T.; Inaoka, D.K.; Kita, K.; Kurisu, G.; Mineki, S.; Harada, S.; Watanabe, Y.; Yoshinari, S.
A conserved lysine residue in the crenarchaea-specific loop is important for the crenarchaeal splicing endonuclease activity (2011), J. Mol. Biol., 405, 92-104.

Cloned(Commentary)

Cloned (Comment)	Organism
expression of His6-tagged wild-type and mutant EndAs in Escherichi coli strain Rosetta 2(DE3)	Aeropyrum pernix

Crystallization (Commentary)

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

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

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

Organism	UniProt	Comment	Textmining
Aeropyrum pernix	Q9YBF1	-	-

Purification (Commentary)

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 and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
additional information	Sulfolobus tokodaii tRNATrp precursor as a substrate	Aeropyrum pernix	?	-	?

Synonyms

Synonyms	Comment	Organism
EndA	-	Aeropyrum pernix
splicing endonuclease	-	Aeropyrum pernix

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
70	-	assay at	Aeropyrum pernix

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
8	-	assay at	Aeropyrum pernix

General Information

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