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Information on EC 2.7.7.B16 - DNA primase
for references in articles please use BRENDA:EC2.7.7.B16preliminary BRENDA-supplied EC number
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2.7.7.B16 DNA primaseSpecify your search results
Word Map
- 2.7.7.B16
-
single-stranded
- helicase
- polymerases
- bacteriophage
- primases
- fork
-
priming
- oligoribonucleotides
- dnab
-
polydt
-
alpha-primase
-
okazaki
-
dna-dependent
- aphidicolin
- ssdna
-
lagging-strand
-
replisome
- alpha-amanitin
-
primer-template
- rntp
-
helicase-primase
-
polymerase-primase
-
rna-primed
-
polymerase-alpha
-
alpha-dna
-
primosome
- primpol
-
template-primer
The enzyme appears in viruses and cellular organisms
Reaction Schemes
+
n
=
dN(pdN)n
+
n
+
n
=
N(pN)n
+
n
Synonyms
dna primase, dnag primase, polptn2, ssoprisl, prisl, pabp41, pabp46, mjpri, pit3 replication protein, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
Mjpri
pIT3 replication protein
PriSL
Sso core primase
SsoPriSL
pIT3 replication protein
the Rep245 protein is the N-terminal domain of the pIT3 replication protein encompassing residues 31245
pIT3 replication protein
the Rep245 protein is the N-terminal domain of the pIT3 replication protein encompassing residues 31245
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
dNTP + n dNTP = dN(pdN)n + n diphosphate
-
-
-
-
NTP + n NTP = N(pN)n + n diphosphate
-
-
-
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ATP + ATP
A(pA)n + n diphosphate
Q5JJ72; Q5JJ73
oligo(dT)30 supports extensive DNA and RNA synthesis. Oligo(dT)30 supports the synthesis of shorter RNA chains than those formed in the presence of oligo(dC)30 as well as the production of higher levels of RNA than DNA
-
-
?
dATP + dATP
dA(pdA)n + n diphosphate
Q5JJ72; Q5JJ73
oligo(dT)30 supports extensive DNA and RNA synthesis
-
-
?
dATP + glycerol
dAMP-glycerol + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
dATP + Tris
dAMP-Tris + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
dCTP + glycerol
dAMP-glycerol + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
dCTP + Tris
dAMP-Tris + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
dGTP + glycerol
dGMP-glycerol + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
dGTP + Tris
dGMP-Tris + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
dNTP + n NTP
(dNTP)n+1 + n diphosphate
Q9V292; Q9V291
DNA primase has comparable affinities for ribonucleotides and deoxyribonucleotides. The Pabp41 subunit alone has no RNA synthesis activity but could synthesize long (up to 3 kb) DNA strands. Addition of the Pabp46 subunit increases the rate of DNA synthesis but decreases the length of the DNA fragments synthesized and confers RNA synthesis capability. DNA primase also displayed DNA polymerase, gapfilling, and strand-displacement activities
-
-
?
dTTP + glycerol
dTMP-glycerol + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
dTTP + Tris
dTMP-Tris + diphosphate
Q5JJ72; Q5JJ73
the products are formed by the p41 catalytic subunit alone and the p41-p46 complex in the absence of a DNA template. They are not formed with preparations containing the catalytically inactive p41 subunit
-
-
?
ATP + n ATP
A(pA)n + n diphosphate
the enzyme synthesizes substantially more products on oligo(dC) with GTP as the substrate than on oligo(dT) with ATP as the substrate
-
-
?
ATP + n ATP
A(pA)n + n diphosphate
the enzyme synthesizes substantially more products on oligo(dC) with GTP as the substrate than on oligo(dT) with ATP as the substrate
-
-
?
dGTP + n dGTP
dG(pdG)n + n diphosphate
the catalytic efficiency of the primase heterodimer for dGMP incorporation is 3.6fold higher than that for GMP incorporation
-
-
?
dGTP + n dGTP
dG(pdG)n + n diphosphate
Q5JJ72; Q5JJ73
oligo(dC)30 supports extensive DNA and RNA synthesis. Of the four homo-oligodeoxynucleotides, 30 nt in length oligo(dC)30 is the most effective template supporting extensive DNA synthesis with dGTP. dGTP incorporation exceeds the level of oligo(dC)30 template added, and the lengths of DNA chains are 100 nt
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
Q9P9H1; Q8U4H7
the p41-p46 primase preferentially uses dNTPs compared to NTPs. DNA strand synthesis is stimulated by the addition of NTPs or by addition of ATP to the same level. ATP is the preferable initiating nucleotide for the p41-p46 complex
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
de novo DNA synthesis is 10 times more effective than RNA synthesis
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
-
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
-
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
the enzyme is capable of template-dependent nucleotide polymerization across discontinuous dsDNA templates. The products are shorter than those obtained on ssDNA templates. It prefers NTP over dNTP in template-dependent nucleotide polymerization. The enzyme synthesizes substantially more products on oligo(dC) with GTP as the substrate than on oligo(dT) with ATP as the substrate
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
the oligonucleotides 20-mer oligo(dA), -(dC), -(dG) and -(dT) are used with varying efficiencies, with oligo(d)A being the poorest substrate and oligo(dC) the most preferred one. The enzyme is capable of utilising both ribonucleotides and deoxyribonucleotides for primer synthesis in the presence of natural, or synthetic, single-stranded DNA. It has a significantly higher affinity for ribonucleotides than for deoxyribonucleotides. In addition to the primase and polymerase activities, the enzyme possesses a template-independent 3'-terminal nucleotidyl transferase activity
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
utilizes poly-pyrimidine single-stranded DNA templates with low efficiency for de novo synthesis of RNA primers. Its synthetic function is specifically activated by thymine-containing synthetic bubble structures that mimic early replication intermediates. The enzyme is able to incorporate dATP on oligo(A)15/poly(dT) but with lower efficiency with respect to ATP. In addition, the Sso DNA primase is not able to elongate oligo(rA)15 when it is not annealed to poly(dT) or to initiate de novo synthesis on poly(dT) with dATP
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
the oligonucleotides 20-mer oligo(dA), -(dC), -(dG) and -(dT) are used with varying efficiencies, with oligo(d)A being the poorest substrate and oligo(dC) the most preferred one. The enzyme is capable of utilising both ribonucleotides and deoxyribonucleotides for primer synthesis in the presence of natural, or synthetic, single-stranded DNA. It has a significantly higher affinity for ribonucleotides than for deoxyribonucleotides. In addition to the primase and polymerase activities, the enzyme possesses a template-independent 3'-terminal nucleotidyl transferase activity
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
utilizes poly-pyrimidine single-stranded DNA templates with low efficiency for de novo synthesis of RNA primers. Its synthetic function is specifically activated by thymine-containing synthetic bubble structures that mimic early replication intermediates. The enzyme is able to incorporate dATP on oligo(A)15/poly(dT) but with lower efficiency with respect to ATP. In addition, the Sso DNA primase is not able to elongate oligo(rA)15 when it is not annealed to poly(dT) or to initiate de novo synthesis on poly(dT) with dATP
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
the enzyme is capable of template-dependent nucleotide polymerization across discontinuous dsDNA templates. The products are shorter than those obtained on ssDNA templates. It prefers NTP over dNTP in template-dependent nucleotide polymerization. The enzyme synthesizes substantially more products on oligo(dC) with GTP as the substrate than on oligo(dT) with ATP as the substrate
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
Q5JJ72; Q5JJ73
the enzyme supports both DNA and RNA synthesis, whereas the p41 subunit alone marginally produces RNA and synthesizes DNA chains that are longer than those formed by the complex. The primase complex preferentially interacts with dNTP rather than ribonucleoside triphosphates and initiates RNA as well as DNA chains de novo. The archaeal primase complex, in contrast to the eukaryote homolog, can initiate DNA chain synthesis in the absence of ribonucleoside triphosphates. DNA primers formed by the archaeal complex can be elongated extensively by the Thermococcus kodakaraensis DNA polymerase (Pol) B, whereas DNA primers formed by the p41 catalytic subunit alone are not
-
-
?
GTP + n GTP
G(pG)n + n diphosphate
the enzyme synthesizes substantially more products on oligo(dC) with GTP as the substrate than on oligo(dT) with ATP as the substrate
-
-
?
GTP + n GTP
G(pG)n + n diphosphate
the catalytic efficiency of the primase heterodimer for dGMP incorporation is 3.6fold higher than that for GMP incorporation
-
-
?
GTP + n GTP
G(pG)n + n diphosphate
Q5JJ72; Q5JJ73
oligo(dC)30 supports extensive DNA and RNA synthesis
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
synthesise oligoribonucleotides on various pyrimidine single-stranded DNA templates poly(dT) and poly(dC). The enzyme is almost completely inactive on a poly(dA) synthetic template
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
synthesise oligoribonucleotides on various pyrimidine single-stranded DNA templates poly(dT) and poly(dC). The enzyme is almost completely inactive on a poly(dA) synthetic template
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
Q9V292; Q9V291
the multifunctional archaeal primase is involved in priming and repair
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
Q9V292; Q9V291
DNA primase has comparable affinities for ribonucleotides and deoxyribonucleotides. The Pabp41 subunit alone has no RNA synthesis activity but could synthesize long (up to 3 kb) DNA strands. Addition of the Pabp46 subunit increases the rate of DNA synthesis but decreases the length of the DNA fragments synthesized and confers RNA synthesis capability. DNA primase also displayed DNA polymerase, gapfilling, and strand-displacement activities
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
Q9P9H1; Q8U4H7
the p41-p46 complex synthesizes the primers, which can be extended in the combination with other replication proteins from Pyrococcus furiosus
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
Q9P9H1; Q8U4H7
the p41-p46 primase preferentially uses dNTPs compared to NTPs. The primase complex synthesizes RNA segments only when NTPs but not dNTPs are added as the substrates. When both dNTPs and NTPs exist at equal concentrations, no RNA products are observed. ATP is the preferable initiating nucleotide for the p41-p46 complex
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the small subunit of the clamp loader replication factor C (RFC) of Sulfolobus solfataricus interacts with both the catalytic and non-catalytic subunits of the primase. The primase-clamp loader replication factor C interaction modulates the activities of both enzymes and therefore may be involved in the regulation of primer synthesis and the transfer of primers to DNA polymerase in archaea
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
de novo synthesis of larger molecular size RNA products when incubated with M13 mp18 single-stranded DNA in the presence of a ribonucleotide mixture. The Rep245 domain is capable of utilizing both ribonucleotides and deoxyribonucleotides for de novo primer synthesis and it synthesizes DNA products up to several kb in length in a template-dependent manner. The Rep245 primase-polymerase domain harbors also a terminal nucleotidyl transferase activity, being able to elongate the 3'-end of synthetic oligonucleotides in a non-templated manner. The Rep245 domain contains the catalytic residues required for both primase and polymerase activities
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the enzyme is capable of template-dependent nucleotide polymerization across discontinuous dsDNA templates. The products are shorter than those obtained on ssDNA templates. It prefers NTP over dNTP in template-dependent nucleotide polymerization. The enzyme synthesizes substantially more products on oligo(dC) with GTP as the substrate than on oligo(dT) with ATP as the substrate
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the oligonucleotides 20-mer oligo(dA), -(dC), -(dG) and -(dT) are used as template with varying efficiencies, with oligo(d)A being the poorest substrate and oligo(dC) the most preferred one. The enzyme is capable of utilising both ribonucleotides and deoxyribonucleotides for primer synthesis in the presence of natural, or synthetic, single-stranded DNA. It has a significantly higher affinity for ribonucleotides than for deoxyribonucleotides. Products with an apparent length of 14 nt and 7 nt are mainly synthesised. In addition to the primase and polymerase activities, the enzyme possesses a template-independent 3'-terminal nucleotidyl transferase activity
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
utilizes poly-pyrimidine single-stranded DNA templates with low efficiency for de novo synthesis of RNA primers. Its synthetic function is specifically activated by thymine-containing synthetic bubble structures that mimic early replication intermediates. The enzyme is able to incorporate dATP on oligo(rA)15/poly(dT) but with lower efficiency with respect to rATP. In addition, the Sso DNA primase is not able to elongate oligo(rA)15 when it is not annealed to poly(dT) or to initiate de novo synthesis on poly(dT) with dATP
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
de novo synthesis of larger molecular size RNA products when incubated with M13 mp18 single-stranded DNA in the presence of a ribonucleotide mixture. The Rep245 domain is capable of utilizing both ribonucleotides and deoxyribonucleotides for de novo primer synthesis and it synthesizes DNA products up to several kb in length in a template-dependent manner. The Rep245 primase-polymerase domain harbors also a terminal nucleotidyl transferase activity, being able to elongate the 3'-end of synthetic oligonucleotides in a non-templated manner. The Rep245 domain contains the catalytic residues required for both primase and polymerase activities
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the oligonucleotides 20-mer oligo(dA), -(dC), -(dG) and -(dT) are used as template with varying efficiencies, with oligo(d)A being the poorest substrate and oligo(dC) the most preferred one. The enzyme is capable of utilising both ribonucleotides and deoxyribonucleotides for primer synthesis in the presence of natural, or synthetic, single-stranded DNA. It has a significantly higher affinity for ribonucleotides than for deoxyribonucleotides. Products with an apparent length of 14 nt and 7 nt are mainly synthesised. In addition to the primase and polymerase activities, the enzyme possesses a template-independent 3'-terminal nucleotidyl transferase activity
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the small subunit of the clamp loader replication factor C (RFC) of Sulfolobus solfataricus interacts with both the catalytic and non-catalytic subunits of the primase. The primase-clamp loader replication factor C interaction modulates the activities of both enzymes and therefore may be involved in the regulation of primer synthesis and the transfer of primers to DNA polymerase in archaea
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
utilizes poly-pyrimidine single-stranded DNA templates with low efficiency for de novo synthesis of RNA primers. Its synthetic function is specifically activated by thymine-containing synthetic bubble structures that mimic early replication intermediates. The enzyme is able to incorporate dATP on oligo(rA)15/poly(dT) but with lower efficiency with respect to rATP. In addition, the Sso DNA primase is not able to elongate oligo(rA)15 when it is not annealed to poly(dT) or to initiate de novo synthesis on poly(dT) with dATP
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the enzyme is capable of template-dependent nucleotide polymerization across discontinuous dsDNA templates. The products are shorter than those obtained on ssDNA templates. It prefers NTP over dNTP in template-dependent nucleotide polymerization. The enzyme synthesizes substantially more products on oligo(dC) with GTP as the substrate than on oligo(dT) with ATP as the substrate
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
Q5JJ72; Q5JJ73
the enzyme supports both DNA and RNA synthesis, whereas the p41 subunit alone marginally produces RNA and synthesizes DNA chains that are longer than those formed by the complex. The primase complex preferentially interacts with dNTP rather than ribonucleoside triphosphates and initiates RNA as well as DNA chains de novo. The archaeal primase complex, in contrast to the eukaryote homolog, can initiate DNA chain synthesis in the absence of ribonucleoside triphosphates. DNA primers formed by the archaeal complex can be elongated extensively by the Thermococcus kodakaraensis DNA polymerase (Pol) B, whereas DNA primers formed by the p41 catalytic subunit alone are not. When M13DNA is used as substrate all labeled rNTPs and dNTPs support RNA and DNA synthesis
-
-
?
additional information
?
-
the enzyme although shows 3'-terminal nucleotidyl-transferase activity
-
-
?
additional information
?
-
-
the enzyme although shows 3'-terminal nucleotidyl-transferase activity
-
-
?
additional information
?
-
-
interaction of Sulfolobus solfataricus DnaG primase (SsoDnaG) with the replicative minichromosome maintenance helicase (SsoMCM) on DNA. The site of interaction is mapped. The complex of SsoDnaG with SsoMCM stimulates the ATPase activity of SsoMCM but leaves the priming activity of SsoDnaG unchanged
-
-
?
additional information
?
-
usage of M13mp18ssDNA as a template
-
-
?
additional information
?
-
-
usage of M13mp18ssDNA as a template
-
-
?
additional information
?
-
the enzyme although shows 3'-terminal nucleotidyl-transferase activity
-
-
?
additional information
?
-
-
primase PolpTN2 is able to prime the synthesis of dsDNA by Taq DNA polymerase and by DNA polymerase PolB encoded by the chromosome of Thermococcus nautili using M13mp18ssDNA as a template, method development and evaluation, overview. Mechanism of template-free synthesis of seeds by PolB and PolpTN2 from dNTP substrates
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
additional information
?
-
-
interaction of Sulfolobus solfataricus DnaG primase (SsoDnaG) with the replicative minichromosome maintenance helicase (SsoMCM) on DNA. The site of interaction is mapped. The complex of SsoDnaG with SsoMCM stimulates the ATPase activity of SsoMCM but leaves the priming activity of SsoDnaG unchanged
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
de novo DNA synthesis is 10 times more effective than RNA synthesis
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
-
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
-
-
-
?
dNTP + n dNTP
dN(pdN)n + n diphosphate
-
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
Q9V292; Q9V291
the multifunctional archaeal primase is involved in priming and repair
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
Q9P9H1; Q8U4H7
the p41-p46 complex synthesizes the primers, which can be extended in the combination with other replication proteins from Pyrococcus furiosus
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the small subunit of the clamp loader replication factor C (RFC) of Sulfolobus solfataricus interacts with both the catalytic and non-catalytic subunits of the primase. The primase-clamp loader replication factor C interaction modulates the activities of both enzymes and therefore may be involved in the regulation of primer synthesis and the transfer of primers to DNA polymerase in archaea
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
-
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
the small subunit of the clamp loader replication factor C (RFC) of Sulfolobus solfataricus interacts with both the catalytic and non-catalytic subunits of the primase. The primase-clamp loader replication factor C interaction modulates the activities of both enzymes and therefore may be involved in the regulation of primer synthesis and the transfer of primers to DNA polymerase in archaea
-
-
?
NTP + n NTP
N(pN)n + n diphosphate
-
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
K+
activity requires divalent cations such Mg2+, Mn2+ or Zn2+, and is additionally stimulated by the monovalent cation K+
Mg2+
Mn2+
Zn2+
additional information
Mg2+
activity requires divalent cations such Mg2+, Mn2+ or Zn2+, and is additionally stimulated by the monovalent cation K+
Mg2+
Q9V292; Q9V291
optimal concentration: 10 mM. Activity is dependent on the presence of Mg2+ and Mn2+. Increased MgCl2 concentrations only slightly enhance NTP incorporation, in contrast to MnCl2
Mg2+
the enzyme is strictly dependent on divalent cations, activating metal preferably used by Rep245 for its DNA polymerase activity is Mg2+ at concentrations between 5 and 10 mM
Mn2+
activity requires divalent cations such Mg2+, Mn2+ or Zn2+, and is additionally stimulated by the monovalent cation K+
Mn2+
Q9V292; Q9V291
optimal concentration: 5 mM. Activity is dependent on the presence of Mg2+ and Mn2+. Increased MgCl2 concentrations only slightly enhance NTP incorporation, in contrast to MnCl2
Mn2+
the enzyme is strictly dependent on divalent cations, with Mn2+ as a cofactor, the DNA polymerase activity of Rep245 is optimal at 12.5 mM and decreases noticeably at increasing amounts of Mn2+
Mn2+
the enzyme requires manganese ions for catalytic activity
Mn2+
-
in a MnCl2-soaked crystal, a Mn2+ is bound to one of the oxygens of the Asp111 side chain
Mn2+
Q5JJ72; Q5JJ73
RNA synthesis with the Thermococcus kodakaraensis primase complex is stimulated about 2fold by the presence of Mn2+, whereas the size of RNA chains is marginally affected. DNA synthesis is slightly inhibited by Mn2+
Zn2+
activity requires divalent cations such Mg2+, Mn2+ or Zn2+, and is additionally stimulated by the monovalent cation K+
Zn2+
Q9P9H1; Q8U4H7
the zinc ion is located within the loop region between beta-strand 6 and the prominent helix alpha4. It is coordinated by Cys 106 and His 108 of the loop and Cys114and Cys117 at the beginning of helix alpha4
Zn2+
contain one zinc ion in the small subunits. Residues Cys116, Cys119, Cys128 and Asp131 coordinate the zinc atom
additional information
Rep245 polymerase activity is strictly dependent on divalent cations (Mg2+ or Mn2+), Zn2+ cations do not support the DNA polymerization activity of Rep245
additional information
-
Rep245 polymerase activity is strictly dependent on divalent cations (Mg2+ or Mn2+), Zn2+ cations do not support the DNA polymerization activity of Rep245
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
dATP
Q5JJ72; Q5JJ73
synthesis of the short RNA chains is inhibited at all levels of dATP added, and the size of oligo(rA) chains formed and the amount of ATP incorporated are reduced
dGTP
Q5JJ72; Q5JJ73
dGTP at a molar ratio of dGTP to dATP or rATP of 10:1 inhibits both DNA and RNA synthesis. Lower molar ratios of dGTP:rATP (0.1:1) inhibit ATP incorporation by 91%, whereas dATP incorporation is reduced by 8%
GTP
Q5JJ72; Q5JJ73
dGTP at a molar ratio of dGTP to dATP or rATP of 10:1 reduces dATP incorporation by 43% and ATP incorporation by 92%
Mn2+
Q5JJ72; Q5JJ73
RNA synthesis with the Thermococcus kodakaraensis primase complex is stimulated about 2fold by the presence of Mn2+, whereas the size of RNA chains is marginally affected. DNA synthesis is slightly inhibited by Mn2+
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
ATP
Q5JJ72; Q5JJ73
in the presence of high levels of ATP (ATP:dATP molar ratio of 10:1), dAMP incorporation is stimulated 3-fold, although the size of dAMP-labeled products formed is reduced
additional information
synthetic function is specifically activated by thymine-containing synthetic bubble structures that mimic early replication intermediates
-
additional information
-
synthetic function is specifically activated by thymine-containing synthetic bubble structures that mimic early replication intermediates
-
additional information
PriL but not PriX enhances primer extension by PriS
-
additional information
-
PriL but not PriX enhances primer extension by PriS
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.028
dNTP
pH 8.0, 60°C, in presence of M13 ssDNA as template
0.0032
NTP
recombinant PriSLX, pH 6.5, 55°C
0.025
NTP
pH 8.0, 60°C, in presence of M13 ssDNA as template
additional information
additional information
Q9V292; Q9V291
kinetic parameters for the DNA primase under different metal ion conditions
-
additional information
additional information
-
kinetic parameters for the DNA primase under different metal ion conditions
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0000013
NTP
recombinant PriSL, pH 6.5, 55°C
0.00078
NTP
recombinant PriSX, pH 6.5, 55°C
0.0068
NTP
recombinant PriSLX, pH 6.5, 55°C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
8
the optimal glycineNaOH buffer is at pH 9 (pH 8 at 60°C)
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7 - 8.5
pH 7.0: about 40% of maximal activity, pH 8.5: about 50% of maximal activity
7.5 - 8.5
pH 7.5: about 40% of maximal activity, pH 8.5: about 70% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
60
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
45 - 75
45°C: about 40% of maximal activity, 75°C: about 40% of maximal activity
55 - 70
polymerization reactions on C35 or C34ddC with rGTP. Both the size and the quantity of the products on C35 increase, while synthesis on C34ddC drastically decreases, when the reaction temperature is raised from 55°C to 70°C
68 - 80
-
activity range of template?free synthesis, which is highly temperature-dependent
additional information
-
delayed kinetics of the reaction at low temperature
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
O57934: small subunit, O57935: large subunit
SwissProt
Q5JJ72: small subunit and Q5JJ73: large subunit
Q5JJ72; Q5JJ73
SwissProt
Q9UWW1: large subunit, Q97Z83: small subunit
SwissProt
Q9UWW1: small subunit, Q97Z83: large subunit
SwissProt
subunits PriL, PriS, and PriX; genes priL, priS, and SSO0502
UniProt
Q9UWW1: large subunit, Q97Z83: small subunit
SwissProt
Q9UWW1: small subunit, Q97Z83: large subunit
SwissProt
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
the majority of primases are classified into two major groups, that is, the bacterial DnaG-type primases and the more complex eukaryotic primases. Archaea encode both types of DNA primase, they encode a eukaryotic-type primase comprising a catalytic subunit (PriS) and a noncatalytic subunit (PriL). Identification of a primase noncatalytic subunit, termed PriX, from the hyperthermophilic archaeon Sulfolobus solfataricus. Phylogenomic analysis, overview
metabolism
-
helicase is not essential but accelerates the reaction. HelpTN2 is a helicase of the SF1 family also encoded by pTN2 who enhances the PolpTN2-primed synthesis of dsDNA by PolB using M13mp18 as a template
physiological function
additional information
physiological function
at the initiation of chromosomal DNA replication, DNA primases synthesize short RNA primers, which are subsequently elongated by DNA polymerases
physiological function
Q9V292; Q9V291
the heterodimeric primase complex is necessary for archaeal survival
physiological function
-
DNA primase PolpTN2 and DNA polymerase PolB are essential for synthesis of dsDNA from deoxynucleotide triphosphates by producing an oligonucleotide primer. The first step of template-free DNA synthesis may involve the creation of seeds by PolpTN2 primase, which are subsequently amplified in synergy with PolB. Template?free synthesis is highly temperature?dependent
physiological function
DNA primases play a critical role in the initiation of DNA replication by synthesizing RNA primers on both the leading and lagging strands since DNA polymerases are incapable of de novo DNA synthesis. Product synthesis by PriSL in the presence of PriX, the Sso0502 product, is over an order of magnitude (about 16fold) more efficient than that in its absence. PriX and PriL bind template DNA differently. Both PriSX and PriSLX differ from PriSL in primer synthesis, overview. PriL but not PriX enhances primer extension by PriS
additional information
the catalytic activity of PriS is modulated through concerted interactions with the two noncatalytic subunits in primer synthesis. The catalytic activity of PriS is modulated through concerted interactions with the two noncatalytic subunits in primer synthesis. PriX subunit residues 26-54 are in a flexible region. PriX residues 55-154 fold into a single domain containing 6 helices, which form a compact core stabilized by extensive hydrophobic interactions. The overall structure of the PriX protein is quite unique
additional information
-
the catalytic activity of PriS is modulated through concerted interactions with the two noncatalytic subunits in primer synthesis. The catalytic activity of PriS is modulated through concerted interactions with the two noncatalytic subunits in primer synthesis. PriX subunit residues 26-54 are in a flexible region. PriX residues 55-154 fold into a single domain containing 6 helices, which form a compact core stabilized by extensive hydrophobic interactions. The overall structure of the PriX protein is quite unique
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). PRIS_METJA
Methanocaldococcus jannaschii (strain ATCC 43067 / DSM 2661 / JAL-1 / JCM 10045 / NBRC 100440)
350
0
41802
Swiss-Prot
-
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
29000
x * 29000, Rep245 domain containing the N-terminal domain of the pIT3 replication protein encompassing residues 31245, SDS-PAGE
40772
Q9P9H1; Q8U4H7
1 * 46209 (large subunit Pfup46) + 1 * 40772 (small subunit Pfup41), calculated from sequence
46209
Q9P9H1; Q8U4H7
1 * 46209 (large subunit Pfup46) + 1 * 40772 (small subunit Pfup41), calculated from sequence
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
heterodimer
heterotrimer
archaea encode a eukaryotic-type primase comprising a catalytic subunit, PriS, and a noncatalytic subunit, PriL and PriX. PriX is a diverged homologue of the C-terminal domain of PriL but lacks the iron-sulfur cluster. PriX, PriL and PriS form a stable heterotrimer (PriSLX). Both PriSX and PriSLX show far greater affinity for nucleotide substrates and are substantially more active in primer synthesis than the PriSL heterodimer. PriL, but not PriX, facilitates primer extension by PriS. The catalytic activity of PriS is modulated through concerted interactions with the two noncatalytic subunits in primer synthesis. PriX subunit residues 26-54 are in a flexible region. PriX residues 55-154 fold into a single domain containing 6 helices, which form a compact core stabilized by extensive hydrophobic interactions. The overall structure of the PriX protein is quite unique, sequence and three-dimensional structure comparisons, overview
additional information
?
x * 29000, Rep245 domain containing the N-terminal domain of the pIT3 replication protein encompassing residues 31245, SDS-PAGE
?
-
x * 29000, Rep245 domain containing the N-terminal domain of the pIT3 replication protein encompassing residues 31245, SDS-PAGE
-
heterodimer
Q9P9H1; Q8U4H7
1 * 46209 (large subunit Pfup46) + 1 * 40772 (small subunit Pfup41), calculated from sequence
additional information
Q9P9H1; Q8U4H7
the Pfup46 protein increases the affinity of the primase to DNA by forming a complex with the catalytic Pfup41 subunit
additional information
-
the Pfup46 protein increases the affinity of the primase to DNA by forming a complex with the catalytic Pfup41 subunit
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
vapor diffusion in sitting drops at 20°C. Crystal structure of the catalytic primase subunit, 2.3 A resolution
Q9P9H1; Q8U4H7
cocrystalization with uridine 5'-triphosphate allowing confirmation of the location of the active site. Construction of a model between the DNA primase and a primer/template DNA based on the complex structure for the primer synthesis
crystallographic studies of of the N-terminal domain (NTD) of PriL (PriLNTD; residues 1222) that bind to PriS, 2.9 A resolution
hanging-drop vapor diffusion method at 20°C, with polyethylene glycol 8000 as the precipitant. The crystals belong to the P3(2)21 with unit-cell parameters a = b = 77.8, c = 129.6 A, and alpha = beta = 90°, gamma = 120°. Crystals of the selenomethionine derivative are obtained by means of a cross-seeding method using native crystals. The data for the native and selenomethionine-substituted crystals are collected to 1.8 and 2.2 A resolution
hanging drop vapor diffusion at 18°C, the structure provides the first three-dimensional description of the large subunit and its interaction with the small subunit
purified recombinant PriX deletion mutant 26-154, X-ray diffraction strutcure determination and analysis at 1.95 A resolution, crystallization of the full-length PriX is unsuccessful
structure-function analysis of the pRN1 primase-polymerase domain. The crystal structure shows a central depression lined by conserved residues. Mutations on one side of the depression reduce DNA affinity. On the opposite side of the depression cluster three acidic residues and a histidine, which are required for primase and DNA polymerase activity. One acidic residue binds a manganese ion, suggestive of a metal-dependent catalytic mechanism. The structure does not show any similarity to DNA polymerases, but is distantly related to archaeal and eukaryotic primases, with corresponding active-site residues
-
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Y155A/Y156A/I157A
mutation reduces PriS binding 1000fold
D101A/D103A
the catalyric site mutant enzyme does not exhibit any detectable catalytic activity, even when higher concentrations of enzyme are utilised. The result demonstrate that the terminal transferase-like activity of PriSL is dependent on the catalytic activity of the primase. D101 and D103 are necessary for PriSL catalytic activity
F164E(PriS)
mutantion in small subunitPriS shows considerably weakened subunit association
F164G/I199K(PriS)_F142E/L163E(PriL)
double mutations of F164G/I199K in small subunit PriS and F142E/L163E in large subunit PriL abolishes the PriS-PriL interaction
G165I(PriS)
mutantion in small subunitPriS shows partial destabilization of the complex
N175A/R176
the affinity of DNA-binding site mutant for NTPs is approximately tenfold lower than that of the wild-type primase and that its enzymatic capability is diminished. Therefore, the mutation of N175 and R176 does not alter the DNA binding properties of the primase but modifies its affinity for free NTPs
R84A/R85A(PriL)
mutation in large subunit shows a marked reduction in the size and amount of RNA product synthesized
D101A/D103A
-
the catalyric site mutant enzyme does not exhibit any detectable catalytic activity, even when higher concentrations of enzyme are utilised. The result demonstrate that the terminal transferase-like activity of PriSL is dependent on the catalytic activity of the primase. D101 and D103 are necessary for PriSL catalytic activity
-
F164E(PriS)
-
mutantion in small subunitPriS shows considerably weakened subunit association
-
F164G/I199K(PriS)_F142E/L163E(PriL)
-
double mutations of F164G/I199K in small subunit PriS and F142E/L163E in large subunit PriL abolishes the PriS-PriL interaction
-
G165I(PriS)
-
mutantion in small subunitPriS shows partial destabilization of the complex
-
N175A/R176
-
the affinity of DNA-binding site mutant for NTPs is approximately tenfold lower than that of the wild-type primase and that its enzymatic capability is diminished. Therefore, the mutation of N175 and R176 does not alter the DNA binding properties of the primase but modifies its affinity for free NTPs
-
R84A/R85A(PriL)
-
mutation in large subunit shows a marked reduction in the size and amount of RNA product synthesized
-
additional information
additional information
mutant primase lacking the zinc-binding motifs has higher activity compared with the wild-type enzyme, as well as a bias toward shorter products of 30100 nucleotides
additional information
generation of a PriX deletion mutant that contains amino acid residues 26-154 for crystallization purposes
additional information
-
generation of a PriX deletion mutant that contains amino acid residues 26-154 for crystallization purposes
additional information
-
mutant primase lacking the zinc-binding motifs has higher activity compared with the wild-type enzyme, as well as a bias toward shorter products of 30100 nucleotides
-
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
80
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified following overexpression in Escherichia coli
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
cloning of the gene for the p58-like protein (gene product: Pfup46), expression in Escherichia coli
Q9P9H1; Q8U4H7
expression in Escherichia coli
genes priL, priS, and SSO0502, DNA and amino acid sequence determination and analysis of PriX, phylogenetic analysis and tree, recombinant expression of a deletion mutant PriX protein containing amino acid residues 26-154
hexa-histidine tagged enzyme is expressed in Escherichia coli
overexpression in Escherichia coli as a fusion protein with a hexa-histidine tag
purified following overexpression in Escherichia coli
small and large subunit are co-expressed in Escherichia coli as hexa-Histidine tagged proteins
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Matsui, E.; Nishio, M.; Yokoyama, H.; Harata, K.; Darnis, S.; Matsui, I.
Distinct domain functions regulating de novo DNA synthesis of thermostable DNA primase from hyperthermophile Pyrococcus horikoshii
Biochemistry
42
14968-14976
2003
Pyrococcus horikoshii (O57934 and O57935), Pyrococcus horikoshii
Ito, N.; Matsui, I.; Matsui, E.
Molecular basis for the subunit assembly of the primase from an archaeon Pyrococcus horikoshii
FEBS J.
274
1340-1351
2007
Pyrococcus horikoshii (O57934 and O57935)
Prato, S.; Vitale, R.M.; Contursi, P.; Lipps, G.; Saviano, M.; Rossi, M.; Bartolucci, S.
Molecular modeling and functional characterization of the monomeric primase-polymerase domain from the Sulfolobus solfataricus plasmid pIT3
FEBS J.
275
4389-4402
2008
Saccharolobus solfataricus (Q9UWW1 and Q97Z83), Saccharolobus solfataricus, Saccharolobus solfataricus P2 (Q9UWW1 and Q97Z83)
Ito, N.; Nureki, O.; Shirouzu, M.; Yokoyama, S.; Hanaoka, F.
Crystal structure of the Pyrococcus horikoshii DNA primase-UTP complex: implications for the mechanism of primer synthesis
Genes Cells
8
913-923
2003
Pyrococcus horikoshii (O57934 and O57935), Pyrococcus horikoshii
Ito, N.; Nureki, O.; Shirouzu, M.; Yokoyama, S.; Hanaoka, F.
Crystallization and preliminary X-ray analysis of a DNA primase from hyperthermophilic archaeon Pyrococcus horikoshii
J. Biochem.
130
727-730
2001
Pyrococcus horikoshii (O57934 and O57935), Pyrococcus horikoshii
Liu, L.; Komori, K.; Ishino, S.; Bocquier, A.A.; Cann, I.K.; Kohda, D.; Ishino, Y.
The archaeal DNA primase: biochemical characterization of the p41-p46 complex from Pyrococcus furiosus
J. Biol. Chem.
276
45484-45490
2001
Pyrococcus furiosus (Q9P9H1 and Q8U4H7), Pyrococcus furiosus
Chemnitz Galal, W.; Pan, M.; Kelman, Z.; Hurwitz, J.
Characterization of DNA primase complex isolated from the archaeon, Thermococcus kodakaraensis
J. Biol. Chem.
287
16209-16219
2012
Thermococcus kodakarensis (Q5JJ72 and Q5JJ73), Thermococcus kodakarensis
Chemnitz Galal, W.; Pan, M.; Giulian, G.; Yuan, W.; Li, S.; Edwards, J.L.; Marino, J.P.; Kelman, Z.; Hurwitz, J.
Formation of dAMP-glycerol and dAMP-Tris derivatives by Thermococcus kodakaraensis DNA primase
J. Biol. Chem.
287
16220-16229
2012
Thermococcus kodakarensis (Q5JJ72 and Q5JJ73), Thermococcus kodakarensis
Lao-Sirieix, S.H.; Bell, S.D.
The heterodimeric primase of the hyperthermophilic archaeon Sulfolobus solfataricus possesses DNA and RNA primase, polymerase and 3-terminal nucleotidyl transferase activities
J. Mol. Biol.
344
1251-1263
2004
Saccharolobus solfataricus (Q9UWW1 and Q97Z83), Saccharolobus solfataricus, Saccharolobus solfataricus P2 (Q9UWW1 and Q97Z83)
Le Breton, M.; Henneke, G.; Norais, C.; Flament, D.; Myllykallio, H.; Querellou, J.; Raffin, J.P.
The heterodimeric primase from the euryarchaeon Pyrococcus abyssi: a multifunctional enzyme for initiation and repair?
J. Mol. Biol.
374
1172-1185
2007
Pyrococcus abyssi (Q9V292 and Q9V291), Pyrococcus abyssi
Wu, K.; Lai, X.; Guo, X.; Hu, J.; Xiang, X.; Huang, L.
Interplay between primase and replication factor C in the hyperthermophilic archaeon Sulfolobus solfataricus
Mol. Microbiol.
63
826-837
2006
Saccharolobus solfataricus (Q9UWW1 and Q97Z83), Saccharolobus solfataricus P2 (Q9UWW1 and Q97Z83)
Augustin, M.A.; Huber, R.; Kaiser, J.T.
Crystal structure of a DNA-dependent RNA polymerase (DNA primase)
Nat. Struct. Biol.
8
57-61
2001
Pyrococcus furiosus (Q9P9H1 and Q8U4H7)
Lao-Sirieix, S.H.; Nookala, R.K.; Roversi, P.; Bell, S.D.; Pellegrini, L.
Structure of the heterodimeric core primase
Nat. Struct. Mol. Biol.
12
1137-1144
2005
Saccharolobus solfataricus (Q9UWW1 and Q97Z83), Saccharolobus solfataricus P2 (Q9UWW1 and Q97Z83)
Desogus, G.; Onesti, S.; Brick, P.; Rossi, M.; Pisani, F.M.
Identification and characterization of a DNA primase from the hyperthermophilic archaeon Methanococcus jannaschii
Nucleic Acids Res.
27
4444-4450
1999
Methanocaldococcus jannaschii (Q58249), Methanocaldococcus jannaschii, Methanocaldococcus jannaschii JAL-1 (Q58249)
De Falco, M.; Fusco, A.; De Felice, M.; Rossi, M.; Pisani, F.M.
The DNA primase of Sulfolobus solfataricus is activated by substrates containing a thymine-rich bubble and has a 3'-terminal nucleotidyl-transferase activity
Nucleic Acids Res.
32
5223-5230
2004
Saccharolobus solfataricus (Q9UWW1 and Q97Z83), Saccharolobus solfataricus, Saccharolobus solfataricus P2 (Q9UWW1 and Q97Z83)
Hu, J.; Guo, L.; Wu, K.; Liu, B.; Lang, S.; Huang, L.
Template-dependent polymerization across discontinuous templates by the heterodimeric primase from the hyperthermophilic archaeon Sulfolobus solfataricus
Nucleic Acids Res.
40
3470-3483
2012
Saccharolobus solfataricus (Q9UWW1 and Q97Z83), Saccharolobus solfataricus, Saccharolobus solfataricus P2 (Q9UWW1 and Q97Z83)
Bauer, R.J.; Graham, B.W.; Trakselis, M.A.
Novel interaction of the bacterial-like DnaG primase with the MCM helicase in archaea
J. Mol. Biol.
425
1259-1273
2013
Saccharolobus solfataricus
Lipps, G.; Weinzierl, A.O.; von Scheven, G.; Buchen, C.; Cramer, P.
Structure of a bifunctional DNA primase-polymerase
Nat. Struct. Mol. Biol.
11
157-162
2004
Sulfolobus islandicus
Beguin, P.; Gill, S.; Charpin, N.; Forterre, P.
Synergistic template-free synthesis of dsDNA by Thermococcus nautili primase PolpTN2, DNA polymerase PolB, and pTN2 helicase
Extremophiles
19
69-76
2015
Thermococcus nautili
Liu, B.; Ouyang, S.; Makarova, K.S.; Xia, Q.; Zhu, Y.; Li, Z.; Guo, L.; Koonin, E.V.; Liu, Z.J.; Huang, L.
A primase subunit essential for efficient primer synthesis by an archaeal eukaryotic-type primase
Nat. Commun.
6
7300
2015
Saccharolobus solfataricus (Q9UWW1 AND Q97Z83 AND Q97ZS7), Saccharolobus solfataricus
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