DNA Primase (p58)

In the eukaryotic cell, DNA primase initiates DNA replication by the synthesis of small ribonucleotides called primers. The eukaryotic primase is composed of two subunits, p49 and p58, which purify as a complex tightly bound to DNA polymerase alpha. The tight association of primase with DNA polymerase a implicates the DNA polymerase a as the lagging-strand DNA polymerase in replication. One unique property of primases (as well as RNA polymerases) is the ability to synthesize nucleotides de novo on a template by the formation of an initial dinucleotide. Primase initiates synthesis with a triphosphate purine moiety at the 5'-end. Relatively few errors are made during the formation of the dinucleotide, whereas the primase readily misincorporates ribonucleotides during elongation of this dinucleotide. After synthesis of 7-10 ribonucleotides, the primer-template is translocated intramolecularly to the active site of the DNA polymerase alpha subunit. The p49 subunit of DNA primase contains the catalytic active site. Analyses of mutant proteins indicate that residue 104-111 are most critical for primer synthesis and form part of the active part. Alanine substitution in residues Glu105, Asp109, and Asp111 produces protein with no detectable activity in direct primase assays, indicating that these residues may form part of a conserved carboxylic triad also observed in the active sites of DNA polymerases and reverse transcriptases. The human primase subunits are 90% identical in amino acid sequence to the mouse homologs.\n\nApplications: \nSuitable for use in Western Blot. Other applications not tested.\n\nRecommended Dilution:\nWestern Blot: 1-5ug/ml \n\nOptimal dilutions to be determined by the researcher.\n\nPositive Control: \nHeLa cells\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. For long-term storage, store at -20 degrees C. Aliquots are stable for at least 12 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.

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SPECIFICATIONS

Catalog Number

D3948-11

Size

50ug

Applications

WB

Hosts

Rabbit

Reactivities

Hum, Mouse

Form

Supplied as a liquid in PBS, pH 7.5, 0.2% BSA, 0.05% sodium azide, 40% glycerol.

P Type

Pab

Purity

Purified by Protein A affinity chromatography.

Isotype

IgG

References

1. Stadlbauer, F., et al., Eur. J. Biochem. 222: 781 (1994). 2. Cloutier, S., et al., Genomics 43: 398 (1997). 3. Adlear, D.A., et al., Genomics 9: 642 (1991). 4. Shiratori, A., et al., Genomics 44: 251 (1997). 5. Prussak, C.E., et al., JBC 264: 4957 (1989).

Additional Info

Reacts with human and mouse p58 subunit. No reactivity is seen with p49 subunit.