Literature summary for 3.1.21.B2 extracted from

Wang, D.; Miyazono, K.I.; Tanokura, M.
Tetrameric structure of the restriction DNA glycosylase R.PabI in complex with nonspecific double-stranded DNA (2016), Sci. Rep., 6, 35197-35197 .

Search for more literature for 3.1.21.B2

Crystallization (Commentary)

Crystallization (Comment)	Organism
crystallization experiments with the mutant enzyme R32A/E63A-nonspecific dsDNA complex are performed using the sitting-drop vapour-diffusion method at 20°C. The crystals of the mutant enzyme R32A/E63A-nonspecific dsDNA complex are obtained using a reservoir solution of 0.2 M calcium acetate, 0.1 M imidazole (pH 8.0), and 10% PEG 8000.The structure of enzyme in complex with double-stranded DNA without the R.PabI recognition sequence is determined by X-ray crystallography. The 1.9 A resolution structure of the complex shows that the enzyme forms a tetrameric structure to sandwich the double-stranded DNA and the tetrameric structure is stabilized by four salt bridges	Pyrococcus abyssi

Crystallization (Comment)

Organism

crystallization experiments with the mutant enzyme R32A/E63A-nonspecific dsDNA complex are performed using the sitting-drop vapour-diffusion method at 20°C. The crystals of the mutant enzyme R32A/E63A-nonspecific dsDNA complex are obtained using a reservoir solution of 0.2 M calcium acetate, 0.1 M imidazole (pH 8.0), and 10% PEG 8000.The structure of enzyme in complex with double-stranded DNA without the R.PabI recognition sequence is determined by X-ray crystallography. The 1.9 A resolution structure of the complex shows that the enzyme forms a tetrameric structure to sandwich the double-stranded DNA and the tetrameric structure is stabilized by four salt bridges

Pyrococcus abyssi

Protein Variants

Protein Variants	Comment	Organism
R70D	the mutation does not show an effect on the nonspecific dsDNA binding affinity. The mutations would prevent the tetramerization of the enzyme on nonspecific dsDNA due to the electrostatic repression of their side chains	Pyrococcus abyssi
Y68F	the mutant enzyme possesses approximately the same base-specific DNA binding ability as the wild-type enzyme but has reduced DNA glycosylase activity. The mutations would prevent the tetramerization of the enzyme on nonspecific dsDNA due to the electrostatic repression of their side chains	Pyrococcus abyssi
Y68F/D71R	the wild-type enzyme forms a tetramer with nonspecific dsDNA and the tetramerization is prevented by the D71R mutation. The mutations would prevent the tetramerization of the enzyme on nonspecific dsDNA due to the electrostatic repression of their side chains. The Y68F/D71R mutant would also prevent the tetramerization of the enzyme on nonspecific dsDNA due to the steric hindrance of the long side chain of D71R. The DNA glycosylase activity with the 24 bp dsDNA substrate is 74% compared with the mutant enzyme Y68Y (a mutant enzyme that possesses approximately the same base-specific DNA binding ability as the wild-type enzyme but has reduced DNA glycosylase activity). The activity with the 500 bp dsDNA substrate is 34% compared with the mutant enzyme Y68Y. The activity with the 3000 bp dsDNA substrate is 19% compared with the mutant enzyme Y68Y	Pyrococcus abyssi
Y68F/R26A	the DNA glycosylase activity with the 24 bp dsDNA substrate is 65% compared with the mutant enzyme Y68Y (a mutant enzyme that possesses approximately the same base-specific DNA binding ability as the wild-type enzyme but has reduced DNA glycosylase activity). The activity with the 500 bp dsDNA substrate is 55% compared with the mutant enzyme Y68Y. The activity with the 3000 bp dsDNA substrate is 27% compared with the mutant enzyme Y68Y	Pyrococcus abyssi
Y68F/R70D	the DNA glycosylase activity with the 24 bp dsDNA substrate is 127% compared with the mutant enzyme Y68Y (a mutant enzyme that possesses approximately the same base-specific DNA binding ability as the wild-type enzyme but has reduced DNA glycosylase activity). The activity with the 500 bp dsDNA substrate is 76% compared with the mutant enzyme Y68Y. The activity with the 3000 bp dsDNA substrate is 19% compared with the mutant enzyme Y68Y	Pyrococcus abyssi
Y68F/R70D/D71R	the DNA glycosylase activity with the 24 bp dsDNA substrate is 105% compared with the mutant enzyme Y68Y (a mutant enzyme that possesses approximately the same base-specific DNA binding ability as the wild-type enzyme but has reduced DNA glycosylase activity). The activity with the 500 bp dsDNA substrate is 71% compared with the mutant enzyme Y68Y. The activity with the 3000 bp dsDNA substrate is 40% compared with the mutant enzyme Y68Y	Pyrococcus abyssi

Organism

Organism	UniProt	Comment	Textmining
Pyrococcus abyssi	Q9V2B6	-	-

Purification (Commentary)

Purification (Comment)	Organism
-	Pyrococcus abyssi

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
DNA + H2O	the enzyme forms a tetrameric structure to sandwich the double-stranded DNA and the tetrameric structure is stabilized by four salt bridges. The residues that form the salt bridges (R70 and D71) are essential for the enzyme to find the recognition sequence, The enzyme does not recognize 5'-GT-3', 5'-TA-3' and 5'-AC-3' steps in dsDNA except for 5'-GTAC-3'	Pyrococcus abyssi	?	-	?

Subunits

Subunits	Comment	Organism
tetramer	the enzyme forms a tetrameric structure to sandwich the double-stranded DNA and the tetrameric structure is stabilized by four salt bridges	Pyrococcus abyssi

Synonyms

Synonyms	Comment	Organism
PAB0105	-	Pyrococcus abyssi