X-ray crystal structures elucidate the nucleotidyl transfer reaction of transcript initiation using two nucleotides

Area of Science:

• Biochemistry
• Structural Biology
• Molecular Biology

Background:

• Bacteriophage N4 RNA polymerase is a single-subunit enzyme crucial for transcription initiation.
• Understanding the atomic details of transcript initiation is key to deciphering RNA polymerase function.

Purpose of the Study:

• To determine the X-ray crystal structures of pre- and postcatalytic forms of the bacteriophage N4 RNA polymerase initiation complex.
• To visualize the complete process of transcript initiation at the atomic level.

Main Methods:

• X-ray crystallography was used to obtain high-resolution structures.
• Structural analysis of pre- and postcatalytic states of the initiation complex.

Main Results:

• The study provides atomic images of transcript initiation by bacteriophage N4 RNA polymerase.
• Substrate loading induces O-helix conformational changes, dependent on correct base pairing.
• Substrate binding facilitates catalytic metal ion binding, aligning reactive groups for nucleotidyl transfer.

Conclusions:

• Catalytic metal binding is proposed as the final step before nucleotidyl transfer in bacteriophage N4 RNA polymerase.
• This work elucidates the mechanism of transcript initiation in single-subunit RNA polymerases.