Literature summary for 2.7.7.7 extracted from

Netz, D.J.; Stith, C.M.; Stuempfig, M.; Koepf, G.; Vogel, D.; Genau, H.M.; Stodola, J.L.; Lill, R.; Burgers, P.M.; Pierik, A.J.
Eukaryotic DNA polymerases require an iron-sulfur cluster for the formation of active complexes (2012), Nat. Chem. Biol., 8, 125-132.

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of POL3 containing a cleavable N-terminal GST-tag, and of POL31 and POL32 from the galactose-inducible GAL1-10 promoter in protease-deficient strain BJ2168, expression of c-Myc-tagged catalytic subunits Pol1, Pol2, and Pol3 in Escherichia coli strain BL21. Expression of the C-terminal domain of Pol3 (amino acids 982-1097) with a N-terminal HA epitope tag or C-terminally Strep-tagged CTD domains of Pol1, Pol2, Pol3 in Escherichia coli strain HMS174 (DE3) pLysS	Saccharomyces cerevisiae

Cloned (Comment)

Organism

recombinant expression of POL3 containing a cleavable N-terminal GST-tag, and of POL31 and POL32 from the galactose-inducible GAL1-10 promoter in protease-deficient strain BJ2168, expression of c-Myc-tagged catalytic subunits Pol1, Pol2, and Pol3 in Escherichia coli strain BL21. Expression of the C-terminal domain of Pol3 (amino acids 982-1097) with a N-terminal HA epitope tag or C-terminally Strep-tagged CTD domains of Pol1, Pol2, Pol3 in Escherichia coli strain HMS174 (DE3) pLysS

Saccharomyces cerevisiae

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
mitochondrion	-	Saccharomyces cerevisiae	5739	-

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Fe2+	required, in [4Fe-4S] clusters, that are bound to the CysB motif in all yeast B family DNA polymerases, assembly of the essential Fe-S cluster is strictly dependent on the function of mitochondrial Nfs1 and cytosolic Nbp35. The C-terminal domain of the catalytic subunit binds the Fe-S cluster in the CysB motif requiring all Cys residues of motif Cysb	Saccharomyces cerevisiae
Zn2+	required, Zn2+ is bound to the CysA motif in yeast B family DNA polymerases	Saccharomyces cerevisiae

Organism

Organism	UniProt	Comment	Textmining
Saccharomyces cerevisiae	-	-	-
Saccharomyces cerevisiae W303-1A	-	-	-

Purification (Commentary)

Purification (Comment)	Organism
GST-tagged POL3, POL31, and POL32 by glutathione affinity chromatography	Saccharomyces cerevisiae

Subunits

Subunits	Comment	Organism
More	the catalytic subunits Pol1, Pol2 and Pol3 are phylogenetically related, and belong to the class B DNA polymerases	Saccharomyces cerevisiae
tetramer	isozymes pol alpha and epsilon contain subunits Pol1, Pol12, Pri1, and Pri2, Pol2, Dpb2, Dpb3 and Dpb4, respectively	Saccharomyces cerevisiae
trimer	isozyme pol delta contains subunits Pol3, Pol31 and Pol32	Saccharomyces cerevisiae

Synonyms

Synonyms	Comment	Organism
pol alpha	-	Saccharomyces cerevisiae
pol delta	-	Saccharomyces cerevisiae
POl epsilon	-	Saccharomyces cerevisiae

General Information

General Information	Comment	Organism
evolution	the catalytic subunits Pol1, Pol2 and Pol3 or isozymes pol alpha, pol epsilon, and pol delta are phylogenetically related, and belong to the class B DNA polymerases. The B subunits are all essential and share a phosphodiesterase-like and oligosaccharide binding domain. Eukaryotes contain a fourth class B DNA polymerase, Pol zeta, which is the major enzyme responsible for mutagenesis in response to DNA damage	Saccharomyces cerevisiae
malfunction	weakened Fe-S cluster binding efficiency of CysA mutant proteins, caused by a lack of polymerase complex stabilization by Pol31, even though this subunit interacts primarily with the CysB region, overexpression of Pol31 results in a 4 to 8fold higher 55Fe binding to both wild-type and CysA mutant Pol3-CTDs. Depletion of the cysteine desulfurase Nfs1 by growth on glucose of the galactose-regulatable strain Gal-NFS1 almost completely abolishes Fe binding to the polymerases, and depletion of the CIA machinery components Nbp35 and Nar1 in regulatable yeast strains abrogates Fe-S cluster formation on the polymerases	Saccharomyces cerevisiae
additional information	physiological importance of the two different metal cofactors, the [4Fe-4S] cluster in CysB and Zn2+ in CysA, in the stabilization of DNA polymerase interactions with different accessory proteins essential for processive DNA synthesis at the replication fork. CysA is an important determinant for proliferating cell nuclear antigen binding	Saccharomyces cerevisiae