Biology Forums - Study Force

?? Why??

DNA polymerase requires a free 3' OH group, in which an RNA primer provides. The enzyme, DNA polymerase III (think of it as a machine), needs a free 3' OH group so it can start creating a complementary DNA strand, based on the template. In other words, it requires a primer because it can't initiate polymerization by itself (it's like a car running without road to drive on); it needs a preexisting free 3' OH group (from the primer) to which it can add new deoxyribonucleoside triphosphates. It does this via the hydrolysis of phosphate groups from the triphosphates to provide the energy to make covalent bonds, forming a phosphodiester linkages. Unlike DNA polymerase, RNA polymerase doesn't require a primer - it can synthesize RNA (in the form of mRNA) de novo. RNA polymerase also unwinds the DNA double helix itself to make it available for transcription.

RNA is not much different from DNA (virtually same as far as polymerization reaction is concerned). If RNA polymerase can start without any primer ... it seems that there is not much mechanistic problem for DNA polymerase in doing so. Does it has something to do with accuracy? Like, the kind of design required to start polymerization from scratch comes at the cost of fidelity (i.e replication process become more error prone). Or there is a fundamental difference (completely different mechanism) in the way RNA polymerase and DNA polymerase work?
By the way a complementary question, why RNA polymerase has low fidelity?

The enzyme, RNA polymerase and DNA polymerase are different (yet they nearly do the same thing) - this doesn't mean they are identical. RNA polymerase does not require the 3' OH to initiate synthesis of RNA molecules. It's as simple as that. During initiation of DNA replication, DNA nucleotide triphosphates must be attached to a 3' OH of an RNA molecule by DNA polymerase. This process, in itself, removes the terminal two phosphates of the nucleotides. (Remember, DNA molecules at the 5' end have only one phosphate, whereas RNA molecules (such as mRNA) have three phosphates at their 5' end - DNA molecules never do.) If an RNA molecule is subsequently removed, then a single phosphate would remain at the 5' end of the DNA molecule. RNA polymerase does not require the 3' OH to initiate synthesis of RNA molecules. Therefore, the 5' end of a RNA molecule will retain all three of the phosphates from the original nucleotide triphosphate substrate. Also, as mentioned earlier, DNA synthesis needs a primer. A good question to ask is how is a primer produced. An enzyme called primase serves this purpose, by synthesizing a short stretch of RNA (generally from 5 to 15 nucleotides in length). RNA polymerase acts very similarly to primase and is able to synthesize a short primer where needed (i.e. during RNA synthesis).

Because it has no proofreading function due to a lack or reduction of 3'-5'exonuclease proof-reading activity.

I drew an image of DNA being synthesized with a primer at the left (uracil). So to conclude, DNA polymerase needs a primer in order to synthesize new DNA because it needs something to latch on. Once the primer is removed, we're left with a 5' end contain only one phosphate. RNA polymerase doesn't require a primer because it's producing RNA. This also allows it to have 3 phosphates at the end of its 5' end.

Thank you very much Bio_man. It was very helpful.
It answers why by the existing structure the DNA polymerase can't initiate DNA replication (polymerization reaction). And I appreciate the answer.
I am also curious about the evolutionary answer. Why evolution selected DNA polymerases unable of initiating dna replication? At least what is rationale for such an enzyme?

Hiya :hey:

Personally I think RNA polymerase is older than DNA polymerase and that DNA polymerase evolved from it, because if you think about it RNA is a simple structure than DNA and you can create simple RNA-like molecules using the Urey-Stanley Miller 1953 Experiment: https://biology-forums.com/index.php?action=gallery;sa=view&id=1 (https://biology-forums.com/index.php?action=gallery;sa=view&id=1) . Technically they are called Ribozymes... anyways if RNA came first, I would need a mechanism for replication.

I read about this the other day. According to the source, it is believed that both polymerases are evolutionarily related, especially since the addition of ribonucleotides to the RNA transcript has a very similar mechanism to DNA polymerization.

Aspartyl (asp) residues in the RNA polymerase hold onto Mg2+ ions, which will, in turn, coordinate the phosphates of the ribonucleotides. The first Mg2+ will hold onto the \({\alpha}\)-phosphate of the nucleotide triphosphosphates (NTP) to be added. This allows the nucleophilic attack of the 3' OH from the RNA transcript, adding an additional NTP to the chain. The second Mg2+ will hold onto the pyrophosphate of the NTP. The overall reaction equation is:

(NMP)n + NTP --> (NMP)n+1 + PPi

I think it has to do with mutations...
In order to detect a mismatch and correct it, a DNA polymerase needs a certain amount of "double helix". In other words, the chance to introduce mutations and not being able to correct them may be higher when dealing with shorter fragments. An RNA primers solve the problem: even if errors are introduced in the first 5-10 nucleotides, they are removed when the RNA primer is replaced by DNA. RNA polymerase does not have this problem, since the first 5-10 nt are in the 5' UTR.

RNA polymerase doesnt require a primer to initiate transcription. this requires the initiating ribonucleotide to be brought to the active site of RNA polymerase and held stably on the template strand while the second NTP is presented with proper geometry for polymerization reaction to occur. So the RNA polymerase enzyme itself has to makeinteractions with first and second ribonucleotides holding one or both rigidly in correct orientation to allow the chemical attack over incoming NTP. This interaction is provided by the various part of the polymerase including \({\sigma}\) region 3/4 linker.

but RNA pol also requires a 3' OH to polymerize the chain forward...isn't it? So why it people say that it does not require a free 3' OH?