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Literature summary extracted from

  • Lemak, S.; Beloglazova, N.; Nocek, B.; Skarina, T.; Flick, R.; Brown, G.; Popovic, A.; Joachimiak, A.; Savchenko, A.; Yakunin, A.F.
    Toroidal structure and DNA cleavage by the CRISPR-associated [4Fe-4S]-cluster containing Cas4 nuclease SSO0001 from Sulfolobus solfataricus (2013), J. Am. Chem. Soc., 135, 17476-17487.
    View publication on PubMedView publication on EuropePMC

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
3.1.12.1 the crystal structure reveals a decameric toroid with a large 55 A diameter central channel formed by five tightly packed dimers Saccharolobus solfataricus

Protein Variants

EC Number Protein Variants Comment Organism
3.1.12.1 C188A the mutant protein is inactive in both binding and cleavage of ssDNA. It forms various protein aggregates Saccharolobus solfataricus
3.1.12.1 C191A the mutant protein is inactive in both binding and cleavage of ssDNA. It forms various protein aggregates Saccharolobus solfataricus
3.1.12.1 C32A the mutant protein is inactive in both binding and cleavage of ssDNA. It forms various protein aggregates Saccharolobus solfataricus
3.1.12.1 D99A inactive mutant enzyme Saccharolobus solfataricus

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.1.12.1 iron-sulfur centre near the active site, each enzyme protomer contains one [4Fe-4S] cluster, which is important for both maintaining the decameric state and nuclease activity Saccharolobus solfataricus
3.1.12.1 K+ nuclease activity of the enzyme is maximal in the presence of 20-200 mM KCl Saccharolobus solfataricus
3.1.12.1 Mg2+ nuclease activity of the enzyme is maximal in the presence of Mg2+ or Mn2+ (5 mM) Saccharolobus solfataricus
3.1.12.1 Mn2+ nuclease activity of the enzyme is maximal in the presence of Mg2+ or Mn2+ (5 mM). Mn2+ is bound in the active site located inside the internal tunnel Saccharolobus solfataricus
3.1.12.1 additional information Ca2+ does not support the nuclease activity Saccharolobus solfataricus

Organism

EC Number Organism UniProt Comment Textmining
3.1.12.1 Saccharolobus solfataricus Q97TX9
-
-
3.1.12.1 Saccharolobus solfataricus P2 Q97TX9
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.1.12.1
-
Saccharolobus solfataricus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.1.12.1 additional information the enzyme also exhibits ATP-independent DNA unwinding activity Saccharolobus solfataricus ?
-
?
3.1.12.1 additional information the enzyme also exhibits ATP-independent DNA unwinding activity Saccharolobus solfataricus P2 ?
-
?
3.1.12.1 poly-dC10 + H2O
-
Saccharolobus solfataricus cytidine 3'-phosphate + ?
-
?
3.1.12.1 poly-dC10 + H2O
-
Saccharolobus solfataricus P2 cytidine 3'-phosphate + ?
-
?
3.1.12.1 single-stranded oligodeoxyribonucleotide + H2O
-
Saccharolobus solfataricus nucleoside 3'-phosphate + ?
-
?
3.1.12.1 single-stranded oligodeoxyribonucleotide + H2O
-
Saccharolobus solfataricus P2 nucleoside 3'-phosphate + ?
-
?
3.1.12.1 ssDNA + H2O short oligonucleotides (17 nt) are cleaved faster than longer substrates (62 nt). Lack of significant sequence preference toward the substrates containing the Sulfolobus solfataricus protospacer adjacent motif (PAMs) in DNA binding and cleavage reactions in vitro. The rates of cleavage of ssRNA and dsDNA with blunt ends by the enzyme are approximately 20 times and 200 times lower, respectively compared to ssDNA Saccharolobus solfataricus nucleoside 3'-phosphate + ?
-
?
3.1.12.1 ssDNA + H2O short oligonucleotides (17 nt) are cleaved faster than longer substrates (62 nt). Lack of significant sequence preference toward the substrates containing the Sulfolobus solfataricus protospacer adjacent motif (PAMs) in DNA binding and cleavage reactions in vitro. The rates of cleavage of ssRNA and dsDNA with blunt ends by the enzyme are approximately 20 times and 200 times lower, respectively compared to ssDNA Saccharolobus solfataricus P2 nucleoside 3'-phosphate + ?
-
?

Subunits

EC Number Subunits Comment Organism
3.1.12.1 decamer crystallized at room temperature using the sitting drop vapour diffusion method. The crystal structure reveals a decameric toroid with a large 55 A diameter central channel formed by five tightly packed dimers Saccharolobus solfataricus

Synonyms

EC Number Synonyms Comment Organism
3.1.12.1 Cas4
-
Saccharolobus solfataricus
3.1.12.1 SSO0001 locus name Saccharolobus solfataricus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.1.12.1 65 70
-
Saccharolobus solfataricus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.1.12.1 9 10
-
Saccharolobus solfataricus

General Information

EC Number General Information Comment Organism
3.1.12.1 physiological function a potential functional role of the enzyme and other Cas4 proteins in CRISPR (Clusters of Regularly interspaced Palindromic Repeats) immunity is based on the presence of 5' to 3' exonuclease and DNA unwinding activities, which produce 3'-ssDNA overhangs as potential intermediates in the process of new spacer addition. The toroidal structures of the enzyme can serve as sliding clamps for other Cas proteins Saccharolobus solfataricus