DNA polymerase III

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The DNA polymerase III is an enzyme which is the synthesis of DNA from deoxyribonucleotides catalyzed on a DNA template. It is a protein complex . The holoenzyme plays the most important role in prokaryotic DNA replication. The most important features are its many subunits and the very high catalytic effect , accuracy and process activity (ability of an enzyme to catalyze many reactions in succession without losing the substrate ). In addition to DNA polymerase III, two other DNA polymerases are also known in prokaryotes.

function

The replication process with primase, helicase, primer and DNA polymerase III.

DNA polymerase III forms several thousand phosphodiester bonds with its substrate before it leaves it. In this way she can hold the template and only release it again after the replication is complete. Furthermore, the DNA polymerase III has a strong catalytic effect , which enables it to add 1000 nucleotides per second . This strong catalyst performance is explained by the fact that they are not from the substrate solve needs such as DNA polymerase I . The new strand grows in a 5 '→ 3' direction. From a chemical point of view, the terminal 3'- OH of the DNA strand undergoes a nucleophilic attack on the 5'- phosphate of the dNTP, with pyrophosphate being split off. The DNA polymerase needs a free 3'-hydroxyl group, i.e. a primer , in order to attach nucleotides to it. It is also possible for DNA polymerase III to proofread 3 '→ 5' and to replace incorrectly incorporated nucleotides via exonuclease activity .

construction

Schematic structure of DNA polymerase III with its subunits

The holoenzyme is compared with the DNA polymerase I, an order of magnitude more difficult, and the molecular weight is close to 900  kDa . The enzyme is designed as an asymmetric dimer to replicate both parent strands in the same place at the same time. The asymmetry arises from the fact that leading and secondary strands are synthesized differently.

The α subunit is the polymerase and the ε is the 3 '→ 5' exonuclease for proofreading . Both are catalytically active but not processive, this is done by the subunits β and τ. The processivity can be explained by the spatial structure of β. This subunit forms a clamp through which the DNA double strand slides and does not have to separate from the substrate.

Replication

The ATP-powered helicase unwinds the DNA double strand and enables both strands to be used as templates. SSB proteins hold the strands apart for the period of replication. The leading and secondary strands are synthesized at the same time, but not in the same way. The holoenzyme begins with the lead strand in which the primer set by the primase is bound and synthesizes it continuously. The subsequent strand is synthesized in a more complicated way because it runs from 5 '→ 3' and a synthesis cannot proceed from 3 '→ 5'. Hence it is synthesized in fragments, resulting in many individual 5'-3 'syntheses. The fragments are also called Okazaki fragments . A fragment is always a part of the single DNA strand that is wound in a loop, which is in the active center of the α-subunit and runs in the same direction as the leading strand. After approx. 1000 nucleotides, the following strand is released and the next loop is led into the active center, for which the primase again sets a primer. The RNA residues of the primers are then removed by DNA polymerase I, since DNA polymerase III lacks the exonuclease function 5 '→ 3'. The resulting gaps are filled by the slower DNA polymerase I and linked by the ligase .

swell

  • Jeremy M. Berg, John L. Tymoczko, Lubert Stryer: Biochemistry. Berlin / Heidelberg 2003, ISBN 3-8274-1303-6 .
  • T. Michael Madigan, M. John Martinko, Jack Parker: Biology of Microorganisms. London 2003, ISBN 0-13-066271-2 .