Primase

In bacteria, primase binds to the DNA helicase forming a complex called the primosome. Primase is activated by the helicase where it then synthesizes a short RNA primer approximately 11 ±1 nucleotides long, to which new nucleotides can be added by DNA polymerase. Archaeal and eukaryote primases are heterodimeric proteins with one large regulatory and one minuscule catalytic subunit. The RNA segments are first synthesized by primase and then elongated by DNA polymerase. Then the DNA polymerase forms a protein complex with two primase subunits to form the alpha DNA Polymerase primase complex. Primase is one of the most error prone and slow polymerases. Primase also acts as a halting mechanism to prevent the leading strand from outpacing the lagging strand by halting the progression of the replication fork. The rate determining step in primase is when the first phosphodiester bond is formed between two molecules of RNA. The primase-helicase complex is used to unwind dsDNA (double-stranded) and synthesizes the lagging strand using RNA primers The majority of primers synthesized by primase are two to three nucleotides long. == Types ==

There are two main types of primase: DnaG found in most bacteria, and the AEP (Archaeo-Eukaryote Primase) superfamily found in archaean and eukaryotic primases. While bacterial primases (DnaG-type) are composed of a single protein unit (a monomer) and synthesize RNA primers, AEP primases are usually composed of two different primase units (a heterodimer) and synthesize two-part primers with both RNA and DNA components. While functionally similar, the two primase superfamilies evolved independently of each other. DnaG The crystal structure of primase in E. coli with a core containing the DnaG protein was determined in the year 2000. The toprim fold is used for binding regulators and metals. The primase uses a phosphotransfer domain for the transfer coordination of metals, which makes it distinct from other polymerases. AEP Eukaryote and archaeal primases tend to be more similar to each other, in terms of structure and mechanism, than they are to bacterial primases. Archaeal and eukaryote primases are heterodimeric proteins with one large regulatory (human PRIM2, p58) and one small catalytic subunit (human PRIM1, p48/p49). The large subunit contains a N-terminal 4Fe–4S cluster, split out in some archaea as PriX/PriCT. The large subunit is implicated in improving the activity and specificity of the small subunit. For example, removing the part corresponding to the large subunit in a fusion protein PolpTN2 results in a slower enzyme with reverse transcriptase activity. == Multifunctional primases ==

s (NTPs); however, primases with polymerase capabilities also have an affinity for deoxyribonucleotides (dNTPs). Primases with terminal transferase functionality are capable of adding nucleotides to the 3’ end of a DNA strand independently of a template. Other enzymes involved in DNA replication, such as helicases, may also exhibit primase activity. In eukaryotes and archaea Human PrimPol (ccdc111 and in restarting stalled replication forks. PrimPol is actively recruited to damaged sites through its interaction with RPA, an adapter protein that facilitates DNA replication and repair. PriS alone preferentially synthesizes strings of DNA; but in combination with PriL, the large subunit, RNA polymerase activity is increased. In Sulfolobus solfataricus, the primase heterodimer PriSL can act as a primase, polymerase, and terminal transferase. PriSL is thought to initiate primer synthesis with NTPs and then switch to dNTPs. The enzyme can polymerize RNA or DNA chains, with DNA products reaching as long as 7000 nucleotides (7 kb). It is suggested that this dual functionality may be a common feature of archaeal primases. In viruses and plasmids AEP enzymes are widespread, and can be found encoded in mobile genetic elements including virus/phages and plasmids. They either use them as a sole replication protein or in combination with other replication-associated proteins, such as helicases and, less frequently, DNA polymerases. Whereas the presence of AEP in eukaryotic and archaeal viruses is expected in that they mirror their hosts, A great diversity of AEP families has been uncovered in various bacterial plasmids by comparative genomics surveys. Vaccinia virus D5 and HSV Primase are examples of AEP-helicase fusion as well. PolpTN2 is an Archaeal primase found in the TN2 plasmid. A fusion of domains homologous to PriS and PriL, it exhibits both primase and DNA polymerase activity, as well as terminal transferase function. Unlike most primases, PolpTN2 forms primers composed exclusively of dNTPs. Unexpectedly, when the PriL-like domain was truncated, PolpTN2 could also synthesize DNA on the RNA template, i.e., acted as an RNA-dependent DNA polymerase (reverse transcriptase). Even DnaG primases can have extra functions, if given the right domains. The T7 phage gp4 is a DnaG primase-helicase fusion, and performs both functions in replication. == References ==