RNA polymerase II

The eukaryotic polymerase responsible for transcription of protein-coding genes and several noncoding transcripts. Its C-terminal-domain phosphorylation state helps regulate elongation.

The RNA polymerase trigger loop is a mobile protein element near the active site of multisubunit RNA polymerase. In RNA polymerase II, it can close beneath a correctly positioned incoming nucleoside triphosphate and form contacts with its base, sugar and phosphate groups. Its movement links recognition of the substrate with efficient phosphodiester bond formation during transcription.^[ref]

The 2006 structural study compared transcribing complexes with different bound nucleotides. A correct substrate in the addition site was associated with a closed conformation and an extensive interaction network. This gave a physical explanation for why merely entering the active-site region is not sufficient for efficient incorporation: the substrate must also support a productive arrangement of the surrounding polymerase.^[ref]

For an illuminating comparison, imagine two nucleotides that bind detectably but support loop closure with different probabilities. Equal binding abundance would not imply equal incorporation rates, because productive positioning contributes a further step. This is a schematic kinetic argument, not a measurement of a particular pair of rate constants.

The adjacent RNA polymerase bridge helix and the RNA polymerase trigger loop are distinct structural elements. A proposed catalytic role for a loop residue must also be distinguished from what is directly observed in a crystal: geometry can suggest a chemical mechanism, while mutagenesis and kinetics test its consequences.

The RNA polymerase bridge helix is an alpha helix spanning the active site region of RNA polymerase. In RNA polymerase II, it lies near the template DNA and growing RNA, where conformational changes can alter the space available for template positioning during transcription.^[ref]

Researchers soaked transcription elongation complex crystals with alpha-amanitin. The trapped structure showed the entering DNA base above a shifted RNA polymerase bridge helix, with the usual templating position obstructed. The authors interpreted the arrangement as a possible intermediate in movement along DNA.^[ref]

The useful connection is between local protein geometry and which DNA base can be presented to the catalytic machinery. Successive addition cycles require that relationship to change while sequence order remains intact. A trapped conformation is evidence about an accessible arrangement; establishing its timing and contribution to normal movement also requires kinetic measurements. The helix is part of the coupled enzyme, rather than an independently operating motor.

The Mediator complex is a multiprotein coactivator involved in coupling regulatory proteins to RNA-polymerase-II transcription. A requirement for mediator activity can be tested by removal and restoration in a reconstituted transcription system.