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Information on EC 2.7.7.102 - DNA primase AEP
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2.7.7.102 DNA primase AEPIUBMB Comments
The enzyme, which is found in eukaryota and archaea, catalyses the synthesis of short RNA or DNA sequences which are used as primers for EC 2.7.7.7, DNA-directed DNA polymerase.
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The expected taxonomic range for this enzyme is: Eukaryota, Archaea
Reaction Schemes
+
n
=
ssDNA/pppN(pN)n-1 hybrid
+
(n-1)
+
n
=
ssDNA/pppdN(pdN)n-1 hybrid
+
(n-1)
Synonyms
protein orf904, ccdc111, prislx, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
AEP
-
-
-
-
archaeo-eukaryotic primase
PriL
PrimPol
PriS
PriSLX
Sso0079
SSO0557
SSO1048
archaeo-eukaryotic primase
-
-
-
-
PrimPol
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ssDNA + n dNTP = ssDNA/pppdN(pdN)n-1 hybrid + (n-1) diphosphate
(2)
-
-
-
ssDNA + n NTP = ssDNA/pppN(pN)n-1 hybrid + (n-1) diphosphate
(1)
-
-
-
SYSTEMATIC NAME
IUBMB Comments
(deoxy)nucleotide 5'-triphosphate:single-stranded DNA (deoxy)nucleotidyltransferase (DNA or DNA-RNA hybrid synthesizing)
The enzyme, which is found in eukaryota and archaea, catalyses the synthesis of short RNA or DNA sequences which are used as primers for EC 2.7.7.7, DNA-directed DNA polymerase.
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
5'-CTTCTTCTGTGC-3' + n dNTP
5'-CTTCTTCTGTGC-3'/pppdN(pdN)n-1 + (n-1) diphosphate
i.e. minimal substrate which supports synthesis of the full-length primer. The base 3' to the central GTG motif is critical for primer synthesis and the bases 5' of the GTG determine the length of the primer or run-off product
-
-
?
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3' + n dNTP
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3'/pppdN(pdN)n-1 + n-1 diphosphate
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3' + n NTP
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3'/pppN(dN)n-1 + n-1 diphosphate
5'-TTTTTTTTGTGCACTTT + n dNTP
5'-TTTTTTTTGTGCACTTT/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
bacteriophage M13 ssDNA + n NTP
bacteriophage M13 ssDNA/pppN(pN)n-1 hybrid + (n-1) diphosphate
M13 ssDNA + n ATP
M13 ssDNA/pppN(pA)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n dATP
M13 ssDNA/pppdN(pdA)n-1 + (n-1) diphosphate
-
-
-
?
M13mp18 ssDNA + n dNTP
M13mp18 ss DNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
M13mp18 ssDNA + n dNTP
M13mp18 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
-
?
poly(dC)2500 + n GTP
poly(dC)2500/pppG(pG)n-1 + n-1 diphosphate
-
-
-
?
poly(dT)220 + n ATP
poly(dT)220/pppdA(pdA)n-1 + (n-1) diphosphate
-
-
-
?
poly(dT)400 + n ATP
poly(dT)400/pppA(pA)n-1 + n-1 diphosphate
-
-
-
?
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3' + n dNTP
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3'/pppdN(pdN)n-1 + n-1 diphosphate
-
-
-
?
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3' + n dNTP
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3'/pppdN(pdN)n-1 + n-1 diphosphate
-
-
-
?
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3' + n NTP
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3'/pppN(dN)n-1 + n-1 diphosphate
-
-
-
?
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3' + n NTP
5'-FAM-AAAAAAAAAAATCAGCGGACAAAAAAAAAAAA-3'/pppN(dN)n-1 + n-1 diphosphate
-
-
-
?
ATP + 7 dNTP
A(pdN)7 + 7 diphosphate
enzyme synthesizes short (8 nt-long) primers in the presence of single-stranded M13 DNA. The 12-bases-long substrate with the sequence 5'-CTTCTTCTGTGC-3' represents the minimal substrate which supports synthesis of the full-length primer. The base 3' to the central GTG motif is critical for primer synthesis and that the bases 5' of the GTG determine the length of the primer or run-off product but do not appear to be important for the efficiency of primer synthesis
-
-
?
ATP + 7 dNTP
A(pdN)7 + 7 diphosphate
formation of a ribonucleotide primer when ORF904 is incubated with ribonucleotides and single-stranded M13 DNA. Effcient dNTP incorporation is strongly dependent on the presence of ATP. The non-hydrolysable analogues beta,gamma-imino-ATP and beta,gamma-methylene-ATP are active in stimulation of primer formation. alpha,beta-methylene-ATP does not enhance primer formation. The stimulatory effect is also seen for GTP and UTP but not for ADP or dATP. The majority of the primers have a length of eight nucleotides. Protein ORF904 shows ATPase, DNA polymerase and primase activity
-
-
?
bacteriophage M13 ssDNA + n NTP
bacteriophage M13 ssDNA/pppN(pN)n-1 hybrid + (n-1) diphosphate
-
-
-
?
bacteriophage M13 ssDNA + n NTP
bacteriophage M13 ssDNA/pppN(pN)n-1 hybrid + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n dNTP
M13 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n dNTP
M13 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
Q9V292; Q9V291
-
-
-
?
M13 ssDNA + n dNTP
M13 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n dNTP
M13 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n dNTP
M13 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n dNTP
M13 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n dNTP
M13 ssDNA/pppdN(pdN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n NTP
M13 ssDNA/pppN(pN)n-1 + (n-1) diphosphate
Q9V292; Q9V291
-
-
-
?
M13 ssDNA + n NTP
M13 ssDNA/pppN(pN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n NTP
M13 ssDNA/pppN(pN)n-1 + (n-1) diphosphate
-
-
-
?
M13 ssDNA + n NTP
M13 ssDNA/pppN(pN)n-1 + (n-1) diphosphate
-
-
-
?
NTP + 7 dNTP
N(pdN)7 + 7 diphosphate
primases are specialized DNA-dependent RNA polymerases that synthesize a short oligoribonucleotide complementary to single-stranded template DNA. In the context of cellular DNA replication, primases are indispensable since DNA polymerases are not able to start DNA polymerization de novo
-
-
?
NTP + 7 dNTP
N(pdN)7 + 7 diphosphate
formation of a ribonucleotide primer when ORF904 is incubated with ribonucleotides and single-stranded M13 DNA. Effcient dNTP incorporation is strongly dependent on the presence of ATP. The non-hydrolysable analogues beta,gamma-imino-ATP and beta,gamma-methylene-ATP are active in stimulation of primer formation. alpha,beta-methylene-ATP does not enhance primer formation. The stimulatory effect is also seen for GTP and UTP but not for ADP or dATP. The majority of the primers have a length of eight nucleotides. Protein ORF904 shows ATPase, DNA polymerase and primase activity
-
-
?
NTP + 7 dNTP
N(pdN)7 + 7 diphosphate
the enzyme synthesizes a mixed primer consisting of a single ribonucleotide at the 5' end followed by seven deoxynucleotides. Ribonucleotides and deoxynucleotides are strictly required at the respective positions within the primer. The primase can initiate primer synthesis with an ATP regardless of the base 5' of the GTG motif and then proceed to synthesize a primer made up of dNTPs. The primase activity is highly sequence-specific and requires the trinucleotide motif GTG in the template. Primer synthesis starts outside of the recognition motif, immediately 5' to the recognition motif
-
-
?
phage X174 ssDNA + n NTP
phage X174 ssDNA/pppN(pN)n-1 + (n-1) diphosphate
-
-
-
?
phage X174 ssDNA + n NTP
phage X174 ssDNA/pppN(pN)n-1 + (n-1) diphosphate
-
-
-
?
ssDNA + n dNTP
ssDNA/pppdN(pdN)n-1 hybrid + (n-1) diphosphate
primases have a fundamental role in DNA replication. They synthesize a primer that is then extended by DNA polymerases
-
-
?
ssDNA + n dNTP
ssDNA/pppdN(pdN)n-1 hybrid + (n-1) diphosphate
a small helical bundle prepares primer synthesis by binding two nucleotides that enhance sequence-specific recognition of the DNA template. Archaeoeukaryotic primases require for synthesis a catalytic and an accessory domain. For the pRN1 archaeal primase, this domain is a 115-amino acid helix bundle domain (HBD). Only the HBD binds the DNA template. DNA binding becomes sequence-specific after a major allosteric change in the HBD, triggered by the binding of two nucleotide triphosphates
-
-
?
additional information
?
-
a template oligonucleotide in which sequence GTCC is flanked by thymine residues is also recognized as substrate, with preferential formation of initiating dinucleotides 5'-A-dG-3' or 5'-dA-dG-3'
-
-
?
additional information
?
-
no substrate: poly(A). In the pH range 6.0-7.6, main products are oilgoribonucleotides of 7 and 14 bases. At pH 8.1 and pH 8.5, longer products are also detected
-
-
?
additional information
?
-
enzyme can synthesize DNA, RNA, and mixed DNA-RNA primers in vitro. At physiologically relevant ratios of substrate, the primase is almost exclusively a RNA synthetic enzyme. There is little discrimination in the initiation site for ATP over dATP, but the inclusion of dATP results in a general decrease of primer length
-
-
?
additional information
?
-
enzyme can synthesize DNA, RNA, and mixed DNA-RNA primers in vitro. At physiologically relevant ratios of substrate, the primase is almost exclusively a RNA synthetic enzyme. There is little discrimination in the initiation site for ATP over dATP, but the inclusion of dATP results in a general decrease of primer length
-
-
?
additional information
?
-
multifunctional replication protein with primase, DNA polymerase and helicase activity. The minimal region required for primase activity encompasses amino-acid residues 40370
-
-
?
additional information
?
-
homopolymers as substrates do not yield detectable primase activity
-
-
?
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
NTP + 7 dNTP
N(pdN)7 + 7 diphosphate
primases are specialized DNA-dependent RNA polymerases that synthesize a short oligoribonucleotide complementary to single-stranded template DNA. In the context of cellular DNA replication, primases are indispensable since DNA polymerases are not able to start DNA polymerization de novo
-
-
?
ssDNA + n dNTP
ssDNA/pppdN(pdN)n-1 hybrid + (n-1) diphosphate
primases have a fundamental role in DNA replication. They synthesize a primer that is then extended by DNA polymerases
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
[4Fe-4S]-center
cluster is buried deeply within the protein core
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
K+
5fold and 2fold activation on substrates poly(dT) and poly(dC), respectively, at 20 mM
Mg2+
Mn2+
Zn2+
Mg2+
or Mn2+, required. Optimum concentration 10 mM
Mn2+
or Mg2+, required. Optimum concentration 11 mM
Zn2+
sequence contains a zinc-finger stem. The zinc ion is tightly bound
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
KCl
in the presence of Mg2+, primase activity is inhibited almost 4fold by increasing the KCl concentration from 15 mM to 150 mM. In the presence of Mn2+, KCl stimulates RNA synthesis
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
KCl
in the presence of Mg2+, primase activity is inhibited almost 4fold by increasing the KCl concentration from 15 mM to 150 mM. In the presence of Mn2+, KCl stimulates RNA synthesis
Mn2+
significantly enhances the binding of nucleotide to primase,which correlates with higher catalytic efficiency in vitro
ATP
ATP binding enhances sequence-specific DNA recognition for primase anchoring
ATP
ATP-induced structural switch that mediates specific DNA recognition in an archaeoeukaryotic primase
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.2
5'-CTTCTTCTGTGC-3'
50°C, pH 7.5, synthesis of full-length primer
additional information
additional information
pH 7.5, 50°C, kinetic properties of the primase activity
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.1
5'-CTTCTTCTGTGC-3'
50°C, pH 7.5, synthesis of full-length primer
additional information
additional information
pH 7.5, 50°C, kinetic properties of the primase activity
-
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.014
5'-CTTCTTCTGTGC-3'
50°C, pH 7.5, synthesis of full-length primer
additional information
additional information
pH 7.5, 50°C, kinetic properties of the primase activity
-
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
Q9V292 i.e. small subunit PriS, Q9V291 i.e. large subunit PriL
Q9V292; Q9V291
SwissProt
Saccharolobus solfataricus DSM 16170
Q97Z83 i.e. small subunit PriS, Q9UWW1 i.e. large subunit PriL
UniProt
P49642 i.e. small subunit Prim1, P09884, i.e. large catalytic subunit
UniProt
P49642 i.e. small subunit Prim1, P49643 i.e. large subunit Prim2
UniProt
P49642 i.e. subunit Prim1, P49643 i.e. large subunit Prim2
UniProt
Q97Z83 i. e. small subunit PriS, Q9UWW1 i.e. large subunit PriL
UniProt
Q97Z83 i.e. small subunit PriS, Q9UWW1 i.e large subunit PriL
UniProt
Q97Z83 i.e. small subunit PriS, Q9UWW1 i.e. large subunit PriL
UniProt
Q9UWW1: DNA primase large subunit PriL, Q97Z83: DNA primase small (catalytic) subunit PriS, Q97ZS7: PriX domain-containing protein
UniProt
Q97Z83 i. e. small subunit PriS, Q9UWW1 i.e. large subunit PriL
UniProt
Q9UWW1: DNA primase large subunit PriL, Q97Z83: DNA primase small (catalytic) subunit PriS, Q97ZS7: PriX domain-containing protein
UniProt
Q97Z83 i.e. small subunit PriS, Q9UWW1 i.e large subunit PriL
UniProt
Q97Z83 i.e. small subunit PriS, Q9UWW1 i.e. large subunit PriL
UniProt
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
physiological function
primases are specialized DNA-dependent RNA polymerases that synthesize a short oligoribonucleotide complementary to single-stranded template DNA. In the context of cellular DNA replication, primases are indispensable since DNA polymerases are not able to start DNA polymerization de novo
physiological function
computational study of the protein sequences and structures of the superfamily of archaeo-eukaryotic primases. Comparison of the enzymes from Pyrococcus furiosus, Sulfolobus islandicus, and others
physiological function
computational study of the protein sequences and structures of the superfamily of archaeo-eukaryotic primases. Comparison of the enzymes from Pyrococcus furiosus, Sulfolobus islandicus, and others
physiological function
does not catalyze by itself the synthesis of short RNA primers but preferentially utilizes deoxynucleotides to synthesize DNA fragments up to several kilobases in length. PriS does not require primers for the synthesis of long DNA strands. PriS interacts with replication protein A
physiological function
isoform PRIMPOL gene silencing or ablation impairs mitochondrial DNA replication. PrimPol is proposed to facilitate replication fork progression by acting as a translesion DNA polymerase or as a specific DNA primase reinitiating downstream of lesions that block synthesis during both mitochondrial and nuclear DNA replication
physiological function
-
primase/polymerase PolpTN2 is encoded by the pTN2 plasmid and exhibits primase, polymerase and nucleotidyl transferase activities. It specifically incorporates dNTPs, to the exclusion of rNTPs. PolpTN2 can efficiently prime DNA synthesis by PolB DNA polymerase. The N-terminal PriS-like domain of PolpTN2 exhibits all activities of the full-length enzyme but is much less efficient in priming cellular DNA polymerases. The N-terminal domain possesses reverse transcriptase activity
physiological function
PriSL is capable of utilising both ribonucleotides and deoxyribonucleotides for primer synthesis in the presence of natural, or synthetic, single-stranded DNA. Products range from dinucleotides to DNA molecules in excess of 7 kb and RNA up to 1 kb in length. PriSL has a significantly higher affinity for ribonucleotides than for deoxyribonucleotides
physiological function
replication protein A directly interacts with primase subunit PrimS
physiological function
the C-terminal domain of the large subunit Prim2 plays a major role in template-primer binding and also defines the elements of the DNA template and the RNA primer that interact with the domain. The interaction with a template-primer involving the terminal 5'-triphosphate of RNA and the 3'-overhang of DNA results in a stable complex between the C-terminal domain of Prim2 and the DNA/RNA duplex
physiological function
Q9P9H1; Q8U4H7
the complex between subunits PriS and PriL shows higher DNA binding activity than the catalytic PriS subunit alone. The amount of DNA synthesized by the complex is much more abundant and shorter in length than that by PriS alone. The activity for RNA primer synthesis is not detected with PriS alone but observed using the complex in vitro
physiological function
the enzyme is able to synthesize products far longer than templates in vitro. The long products result from template-dependent polymerization across discontinuous templates, which is initiated through either primer synthesis or terminal transfer, and occurs efficiently on templates containing contiguous dCs. The enzyme is able to promote strand annealing. PriSL catalyzes template-dependent polymerization across discontinuous templates with either dNTPs or rNTPs as the substrates but prefers the latter
physiological function
The minimal region of the protein required for primase activity encompasses amino-acid residues 40-370. The C-terminal part of the minimal region folds into a compact domain with six helices and is stabilized by a disulfide bond. The C-terminal helix of the helix bundle domain is required for primase activity
physiological function
the primase activity synthesizes a mixed primer consisting of a single ribonucleotide at the 5' end followed by seven deoxynucleotides. Ribonucleotides and deoxynucleotides are strictly required at the respective positions within the primer. The primase activity is highly sequence-specific and requires the trinucleotide motif GTG in the template. Primer synthesis starts outside of the recognition motif, immediately 5' to the recognition motif. Non-complementary bases are not incorporated into the primer
physiological function
Q9V292; Q9V291
the small subunit PrimS alone has no RNA synthesis activity but can synthesize up to 3 kb long DNA strands. Addition of the large subunit increases the rate of DNA synthesis but decreases the length of the DNA fragments synthesized and confers RNA synthesis capability. The primase has comparable affinities for ribonucleotides and deoxyribonucleotides. DNA primase also displays DNA polymerase, gapfilling, and strand-displacement activities
physiological function
primases have a fundamental role in DNA replication. They synthesize a primer that is then extended by DNA polymerases
physiological function
the enzyme plays a role in DNA damage tolerance rather than initiation of DNA replication. Because of its ability to carry out translesion synthesis and to bypass template lesions such as thymine dimers, PrimPol is important for re-priming and resolution of stalled replication forks. Physiologically, PrimPol resolves stalled replication forks that occur if DNA lesions such as UV-induced cyclobutane thymine dimers and (6-4) thymine dimers obstruct the replisome function
physiological function
Saccharolobus solfataricus DSM 16170
-
the enzyme is able to synthesize products far longer than templates in vitro. The long products result from template-dependent polymerization across discontinuous templates, which is initiated through either primer synthesis or terminal transfer, and occurs efficiently on templates containing contiguous dCs. The enzyme is able to promote strand annealing. PriSL catalyzes template-dependent polymerization across discontinuous templates with either dNTPs or rNTPs as the substrates but prefers the latter
-
physiological function
-
PriSL is capable of utilising both ribonucleotides and deoxyribonucleotides for primer synthesis in the presence of natural, or synthetic, single-stranded DNA. Products range from dinucleotides to DNA molecules in excess of 7 kb and RNA up to 1 kb in length. PriSL has a significantly higher affinity for ribonucleotides than for deoxyribonucleotides
-
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). PRI1_HUMAN
420
0
49902
Swiss-Prot
other Location (Reliability: 1)
PRI1_MOUSE
417
0
49295
Swiss-Prot
other Location (Reliability: 1)
PRI1_PLAF7
452
0
53489
Swiss-Prot
other Location (Reliability: 1)
PRI1_PLAFK
452
0
53489
Swiss-Prot
other Location (Reliability: 1)
PRI1_SCHPO
Schizosaccharomyces pombe (strain 972 / ATCC 24843)
454
0
52009
Swiss-Prot
other Location (Reliability: 1)
PRIL_PYRHO
Pyrococcus horikoshii (strain ATCC 700860 / DSM 12428 / JCM 9974 / NBRC 100139 / OT-3)
394
0
45949
Swiss-Prot
-
PRI1_YEAST
Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
409
0
47690
Swiss-Prot
Secretory Pathway (Reliability: 3)
PRIS_METJA
Methanocaldococcus jannaschii (strain ATCC 43067 / DSM 2661 / JAL-1 / JCM 10045 / NBRC 100440)
350
0
41802
Swiss-Prot
-
PRIPO_HUMAN
560
0
64412
Swiss-Prot
other Location (Reliability: 5)
PRIS_PYRFU
Pyrococcus furiosus (strain ATCC 43587 / DSM 3638 / JCM 8422 / Vc1)
347
0
40772
Swiss-Prot
-
PDB
SCOP
CATH
UNIPROT
ORGANISM
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
heterotrimer
?
Q9V292; Q9V291
x * 41000, subunit PrimS, plus x * 46000, subunit PrimL, claculated from sequence
?
x * 35700, subunit PriL, plus x * 37600, subunit PriS
heterotrimer
1 * 35734 (subunit PriS) + 1 * 37598 (subunit PriL) and 1 * 18029 (subunit PriX), calculated from sequence. PriX is essential for primer synthesis and is structurally related to the Fe-S cluster domain of eukaryotic PriL. PriX contains a nucleotide-binding site required for primer synthesis. A primase chimera, where PriX is fused to a truncated version of PriL lacking the Fe-S cluster domain retains wild type levels of primer synthesis
heterotrimer
-
1 * 35734 (subunit PriS) + 1 * 37598 (subunit PriL) and 1 * 18029 (subunit PriX), calculated from sequence. PriX is essential for primer synthesis and is structurally related to the Fe-S cluster domain of eukaryotic PriL. PriX contains a nucleotide-binding site required for primer synthesis. A primase chimera, where PriX is fused to a truncated version of PriL lacking the Fe-S cluster domain retains wild type levels of primer synthesis
-
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
strcuture of the highly conserved C-terminal regulatory domain of the large subunit to 1.7 A resolution. The evolutionarily conserved 4Fe-4S cluster is buried deeply within the protein core. DNA binding shows a strong preference for ss/dsDNA junction substrates. The enzyme interacts specifically with the C-terminal domain of the intermediate subunit of replication protein A, RPA32C
structure of small subunit Prim1 at 2.2 A resolution, with citrate in its inactive forms. Dibasic citrate is bound at the nucleotide triphosphate beta, gamma-phosphate binding site through nine hydrogen bonds. The activity of Prim1 is regulated by pH and citrate
structure of the catalytic subunit, with bound UTP and Mn2+. Primase contains the conserved catalytic prim fold domain, with a subdomain different from the archaeal and bacterial primases. Residues S160 and H166 are in direct contact with UTP
structure of the complex Prim1/Prim2, at 2.65 A resolution, and modeling of the protein in complex with NTP and DNA/RNA substrates
crystal structure in complex with UTP. The primase binds the triphosphate moiety of the UTP at the active site, which includes residues Asp95, Asp97, and Asp280, essential for the nucleotidyl transfer reaction. Construction of a model between the DNA primase and a primer/template DNA for the primer synthesis
N-terminal domain (NTD) of PriL, residues 1-222 that bind to small subunit PriS at 2.9 A resolution. The PriL-NTD structure consists of the helix-bundle and twisted-strand domains. The conserved hydrophobic Tyr155-Tyr156-Ile157 region near the flexible region is the PriS-binding site
native and selenomethionine-substituted protein, to 1.8 and 2.2 A resolution, respectively, space group P3(2)21
vapour diffusion in sitting drops at 19°C. X-ray crystal structure of heterotrimeric PriSLX from S. solfataricus at 2.9 A resolution
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
H299D
10fold reduction in affinity for replication protein A
R302
50fold reduction in affinity for replication protein A
Y155A/Y156A/I157A
D101
catalytic site mutant, complete loss of activity
D103
catalytic site mutant, complete loss of activity
D235E
mutation of aspartic acid by glutamic acid in DNA primase small (catalytic) subunit PriS may occur naturally due to a misrepair on the DNA replication and by a substitution of the third nucleotide of the codons GAU and GAC to GAA to GAG, corresponding to aspartic acid and glutamic acid, respectively. The in silico analysis suggests that these mutations in PriL may cause destabilization on its structure interfering with replication mechanisms of Saccharolobus solfataricus. In addition, the mutation may alter the interactions with other molecules, such as salt bridges
D241E
mutation of aspartic acid by glutamic acid in DNA primase small (catalytic) subunit PriS may occur naturally due to a misrepair on the DNA replication and by a substitution of the third nucleotide of the codons GAU and GAC to GAA to GAG, corresponding to aspartic acid and glutamic acid, respectively. The in silico analysis suggests that these mutations in PriL may cause destabilization on its structure interfering with replication mechanisms of Saccharolobus solfataricus. In addition, the mutation may alter the interactions with other molecules, such as salt bridges
D62E
mutation of aspartic acid by glutamic acid in DNA primase small (catalytic) subunit PriS may occur naturally due to a misrepair on the DNA replication and by a substitution of the third nucleotide of the codons GAU and GAC to GAA to GAG, corresponding to aspartic acid and glutamic acid, respectively. The in silico analysis suggests that these mutations in PriL may cause destabilization on its structure interfering with replication mechanisms of Saccharolobus solfataricus. In addition, the mutation may alter the interactions with other molecules, such as salt bridges
N175
residue involved in DNA-binding. Mutant accumulates dinucleotides, no other products are formed
R176
residue involved in DNA binding. Mutant accumulates dinucleotides, no other products are formed
D235E
-
mutation of aspartic acid by glutamic acid in DNA primase small (catalytic) subunit PriS may occur naturally due to a misrepair on the DNA replication and by a substitution of the third nucleotide of the codons GAU and GAC to GAA to GAG, corresponding to aspartic acid and glutamic acid, respectively. The in silico analysis suggests that these mutations in PriL may cause destabilization on its structure interfering with replication mechanisms of Saccharolobus solfataricus. In addition, the mutation may alter the interactions with other molecules, such as salt bridges
-
D241E
-
mutation of aspartic acid by glutamic acid in DNA primase small (catalytic) subunit PriS may occur naturally due to a misrepair on the DNA replication and by a substitution of the third nucleotide of the codons GAU and GAC to GAA to GAG, corresponding to aspartic acid and glutamic acid, respectively. The in silico analysis suggests that these mutations in PriL may cause destabilization on its structure interfering with replication mechanisms of Saccharolobus solfataricus. In addition, the mutation may alter the interactions with other molecules, such as salt bridges
-
D62E
-
mutation of aspartic acid by glutamic acid in DNA primase small (catalytic) subunit PriS may occur naturally due to a misrepair on the DNA replication and by a substitution of the third nucleotide of the codons GAU and GAC to GAA to GAG, corresponding to aspartic acid and glutamic acid, respectively. The in silico analysis suggests that these mutations in PriL may cause destabilization on its structure interfering with replication mechanisms of Saccharolobus solfataricus. In addition, the mutation may alter the interactions with other molecules, such as salt bridges
-
N175
-
residue involved in DNA-binding. Mutant accumulates dinucleotides, no other products are formed
-
R176
-
residue involved in DNA binding. Mutant accumulates dinucleotides, no other products are formed
-
D111A
D171A
mutation severely reduces primase and abolishes polymerase activity while leaving DNA-binding activity unaffected
delC370
deletion mutant retains the strict ATP dependence for primer synthesis
delC526
deletion mutant retains the strict ATP dependence for primer synthesis
E113A
mutation abolishes primase and polymerase activity while leaving DNA-binding activity unaffected
H141A
W314A
W347A
W361A
Y352A
Y367A
additional information
construction of deletion mutants lacking residues of the linker between the C-terminal and N-terminal regions of subunit Prim2 or having an insertion. Deletion of 15 amino acids of the linker decreases activity about 5fold. The enzyme with a longer linker has the same activity as wild type. Deletion of the C-terminus results in complete loss of the ability to initiate primer synthesis
D111A
the mutant protein is deficient and DNA polymerase activity and primase activity, ATPase activity is unaffected
D111A
mutation abolishes primase and polymerase activity while leaving DNA-binding activity unaffected
W314A
primer formation is severely impaired, very small amount of wild-type like products are formed
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
from insoluble recombinant aggregates, solubilization by addition of high concentrations of glycerol, polyethylene glycol, or Mn2+, and by incubation at 60°C
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
coexpression of both subunits in Saccharomyces cerevisiae and Escherichia coli
expression in Escherichia coli
expression in Escherichia coli, independently and coexpression of subunits
Q9V292; Q9V291
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Lipps, G.; Rther, S.; Hart, C.; Krauss, G.
A novel type of replicative enzyme harbouring ATPase, primase and DNA polymerase activity
EMBO J.
22
2516-2525
2003
Sulfolobus islandicus (Q54324), Sulfolobus islandicus
Beck, K.; Lipps, G.
Properties of an unusual DNA primase from an archaeal plasmid
Nucleic Acids Res.
35
5635-5645
2007
Sulfolobus islandicus (Q54324)
Beck, K.; Vannini, A.; Cramer, P.; Lipps, G.
The archaeo-eukaryotic primase of plasmid pRN1 requires a helix bundle domain for faithful primer synthesis
Nucleic Acids Res.
38
6707-6718
2010
Sulfolobus islandicus (Q54324)
Lee, J.; Park, K.; An, J.; Kang, J.; Shen, H.; Wang, J.; Eom, S.
Structural and biochemical insights into inhibition of human primase by citrate
Biochem. Biophys. Res. Commun.
507
383-388
2018
Homo sapiens (P49642 and P09884)
Bocquier, A.; Liu, L.; Cann, I.; Komori, K.; Kohda, D.; Ishino, Y.
Archaeal primase Bridging the gap between RNA and DNA polymerases
Curr. Biol.
11
452-456
2001
Pyrococcus furiosus (Q9P9H1)
Ito, N.; Nureki, O.; Yokoyama, S.; Hanaoka, F.
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 (O57935), Pyrococcus horikoshii DSM 12428 (O57935)
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 (O57935), Pyrococcus horikoshii DSM 12428 (O57935)
Ito, N.; Nureki, O.; 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 (O57935), Pyrococcus horikoshii DSM 12428 (O57935)
Komori, K.; Ishino, Y.
Replication protein A in Pyrococcus furiosus is involved in homologous DNA recombination
J. Biol. Chem.
276
25654-25660
2001
Pyrococcus furiosus (Q9P9H1)
Liu, L.; Komori, K.; Ishino, S.; Bocquier, A.; Cann, I.; 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)
Baranovskiy, A.; Zhang, Y.; Suwa, Y.; Babayeva, N.; Gu, J.; Pavlov, Y.; Tahirov, T.
Crystal structure of the human primase
J. Biol. Chem.
290
5635-5646
2015
Homo sapiens (P49642 and P49643)
Baranovskiy, A.; Zhang, Y.; Suwa, Y.; Gu, J.; Babayeva, N.; Pavlov, Y.; Tahirov, T.
Insight into the human DNA primase interaction with template-primer
J. Biol. Chem.
291
4793-4802
2016
Homo sapiens (P49642 and P49643)
Lao-Sirieix, S.; Bell, S.
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 (Q97Z83 and Q9UWW1), Saccharolobus solfataricus DSM 1617 (Q97Z83 and Q9UWW1)
Le Breton, M.; Henneke, G.; Norais, C.; Flament, D.; Myllykallio, H.; Querellou, J.; Raffin, J.
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)
Zuo, Z.; Rodgers, C.J.; Mikheikin, A.L.; Trakselis, M.A.
Characterization of a functional DnaG-type primase in archaea implications for a dual-primase system
J. Mol. Biol.
397
664-676
2010
Saccharolobus solfataricus (Q97Z83 and Q9UWW1), Saccharolobus solfataricus ATCC 35092 (Q97Z83 and Q9UWW1)
Vaithiyalingam, S.; Arnett, D.R.; Aggarwal, A.; Eichman, B.F.; Fanning, E.; Chazin, W.J.
Insights into eukaryotic primer synthesis from structures of the p48 subunit of human DNA primase
J. Mol. Biol.
426
558-569
2014
Homo sapiens (P49642), Homo sapiens (P49642 and P49643), Saccharomyces cerevisiae (P10363)
Garcia-Gomez, S.; Reyes, A.; Martinez-Jimenez, M.I.; Chocron, E.S.; Mouron, S.; Terrados, G.; Powell, C.; Salido, E.; Mendez, J.; Holt, I.J.; Blanco, L.
PrimPol, an archaic primase/polymerase operating in human cells
Mol. Cell
52
541-553
2013
Homo sapiens (Q96LW4)
Lipps, G.; Weinzierl, A.; Von Scheven, G.; Buchen, C.; Cramer, P.
Structure of a bifunctional DNA primase-polymerase
Nat. Struct. Mol. Biol.
11
157-162
2004
Sulfolobus islandicus (Q54324)
Desogus, G.; Onesti, S.; Brick, P.; Rossi, M.; Pisani, F.
Identification and characterization of a DNA primase from the hyperthermophilic archaeon Methanococcus jannaschii
Nucleic Acids Res.
27
4444-4450
1999
Methanocaldococcus jannaschii (Q58249)
Iyer, L.M.; Koonin, E.V.; Leipe, D.D.; Aravind, L.
Origin and evolution of the archaeo-eukaryotic primase superfamily and related palm-domain proteins structural insights and new members
Nucleic Acids Res.
33
3875-3896
2005
Pyrococcus furiosus (Q9P9H1), Sulfolobus islandicus (Q54324)
Beck, K.; Lipps, G.
Properties of an unusual DNA primase from an archaeal plasmid
Nucleic Acids Res.
35
5635-5645
2007
Sulfolobus islandicus (Q54324)
Beck, K.; Vannini, A.; Cramer, P.; Lipps, G.
The archaeo-eukaryotic primase of plasmid pRN1 requires a helix bundle domain for faithful primer synthesis
Nucleic Acids Res.
38
6707-6718
2010
Sulfolobus islandicus (Q54324)
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 (Q97Z83 and Q9UWW1), Saccharolobus solfataricus DSM 16170 (Q97Z83 and Q9UWW1)
Gill, S.; Krupovic, M.; Desnoues, N.; Beguin, P.; Sezonov, G.; Forterre, P.
A highly divergent archaeo-eukaryotic primase from the Thermococcus nautilus plasmid, pTN2
Nucleic Acids Res.
42
3707-3719
2014
Thermococcus nautili
Yan, J.; Holzer, S.; Pellegrini, L.; Bell, S.D.
An archaeal primase functions as a nanoscale caliper to define primer length
Proc. Natl. Acad. Sci. USA
115
6697-6702
2018
Saccharolobus solfataricus (Q97Z83 and Q9UWW1), Saccharolobus solfataricus DSM 1617 (Q97Z83 and Q9UWW1)
e Souza, E.; Diniz, M.
Prediction of mutations on structure primase of the archaeon Sulfolobus solfataricus
Acta Sci. Biol. Sci.
39
463-467
2017
Saccharolobus solfataricus (Q9UWW1 AND Q97Z83 AND Q97ZS7), Saccharolobus solfataricus ATCC 35092 (Q9UWW1 AND Q97Z83 AND Q97ZS7)
-
Boudet, J.; Devillier, J.C.; Wiegand, T.; Salmon, L.; Meier, B.H.; Lipps, G.; Allain, F.H.
A small helical bundle prepares primer synthesis by binding two nucleotides that enhance sequence-specific recognition of the DNA template
Cell
176
154-166.e13
2019
Sulfolobus islandicus (Q54324)
Walker, M.J.; Varani, G.
An allosteric switch primes sequence-specific DNA recognition
Cell
176
4-6
2019
Sulfolobus islandicus (Q54324)
Bergsch, J.; Allain, F.H.; Lipps, G.
Recent advances in understanding bacterial and archaeoeukaryotic primases
Curr. Opin. Struct. Biol.
59
159-167
2019
Homo sapiens (Q96LW4)
Holzer, S.; Yan, J.; Kilkenny, M.; Bell, S.; Pellegrini, L.
Primer synthesis by a eukaryotic-like archaeal primase is independent of its Fe-S cluster
Nat. Commun.
8
1718
2017
Saccharolobus solfataricus (Q9UWW1 AND Q97Z83 AND Q97ZS7), Saccharolobus solfataricus ATCC 35092 (Q9UWW1 AND Q97Z83 AND Q97ZS7)
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