PDBsum entry 6o6c

Transferase/DNA/RNA

PDB id: 6o6c

Contents

Protein chains

1446 a.a.

1165 a.a.

270 a.a.

215 a.a.

81 a.a.

146 a.a.

115 a.a.

65 a.a.

111 a.a.

46 a.a.

DNA/RNA

Metals

_MG

_ZN ×8

PDB id:

6o6c

Name:

Transferase/DNA/RNA

Title:

RNA polymerase ii elongation complex arrested at a cpd lesion

Structure:

DNA-directed RNA polymerase ii subunit rpb1. Chain: a. Synonym: RNA polymerase ii subunit b1,DNA-directed RNA polymerase iii largest subunit,RNA polymerase ii subunit b220. DNA-directed RNA polymerase ii subunit rpb2. Chain: b. Synonym: RNA polymerase ii subunit 2,b150,DNA-directed RNA polymerase ii 140 kda polypeptide. DNA-directed RNA polymerase ii subunit rpb3.

Source:

Saccharomyces cerevisiae. Baker's yeast. Organism_taxid: 4932. Synthetic: yes. Organism_taxid: 4932

Authors:

I.Lahiri,A.E.Leshziner

Key ref:

I.Lahiri et al. (2019). 3.1 Å structure of yeast RNA polymerase II elongation complex stalled at a cyclobutane pyrimidine dimer lesion solved using streptavidin affinity grids. J Struct Biol, 207, 270-278. PubMed id: 31200019 DOI: 10.1016/j.jsb.2019.06.004

Date:

05-Mar-19 Release date: 26-Jun-19

Protein chain

P04050  (RPB1_YEAST) -  DNA-directed RNA polymerase II subunit RPB1 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 1733 a.a.

Struc: 1446 a.a.*

Protein chain

P08518  (RPB2_YEAST) -  DNA-directed RNA polymerase II subunit RPB2 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 1224 a.a.

Struc: 1165 a.a.

Protein chain

P16370  (RPB3_YEAST) -  DNA-directed RNA polymerase II subunit RPB3 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 318 a.a.

Struc: 270 a.a.

Protein chain

P20434  (RPAB1_YEAST) -  DNA-directed RNA polymerases I, II, and III subunit RPABC1 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 215 a.a.

Struc: 215 a.a.

Protein chain

P20435  (RPAB2_YEAST) -  DNA-directed RNA polymerases I, II, and III subunit RPABC2 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 155 a.a.

Struc: 81 a.a.

Protein chain

P20436  (RPAB3_YEAST) -  DNA-directed RNA polymerases I, II, and III subunit RPABC3 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 146 a.a.

Struc: 146 a.a.

Protein chain

P27999  (RPB9_YEAST) -  DNA-directed RNA polymerase II subunit RPB9 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 122 a.a.

Struc: 115 a.a.

Protein chain

P22139  (RPAB5_YEAST) -  DNA-directed RNA polymerases I, II, and III subunit RPABC5 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 70 a.a.

Struc: 65 a.a.

Protein chain

P38902  (RPB11_YEAST) -  DNA-directed RNA polymerase II subunit RPB11 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 120 a.a.

Struc: 111 a.a.

Protein chain

P40422  (RPAB4_YEAST) -  DNA-directed RNA polymerases I, II, and III subunit RPABC4 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 70 a.a.

Struc: 46 a.a.

* PDB and UniProt seqs differ at 1 residue position (black cross)

DNA/RNA chains

A-U-C-G-A-G-A-G-G 9 bases

G-G-A-G-A-A-G-G-A-G-C-A-G-A-G-C 16 bases

G-C-T-C-T-G-C-T-C-C-T-T-C-T-C-C-TTD-T-C-C-T-C-T-C-G-A-T 27 bases

Enzyme reactions

• Enzyme class: Chains A, B: E.C.2.7.7.6 - DNA-directed Rna polymerase.
• Reaction: RNA(n) + a ribonucleoside 5'-triphosphate = RNA(n+1) + diphosphate

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

Added reference

DOI no: 10.1016/j.jsb.2019.06.004

J Struct Biol 207:270-278 (2019)

PubMed id: 31200019

3.1 Å structure of yeast RNA polymerase II elongation complex stalled at a cyclobutane pyrimidine dimer lesion solved using streptavidin affinity grids.

I.Lahiri, J.Xu, B.G.Han, J.Oh, D.Wang, F.DiMaio, A.E.Leschziner.

ABSTRACT

Despite significant advances in all aspects of single particle cryo-electron microscopy (cryo-EM), specimen preparation still remains a challenge. During sample preparation, macromolecules interact with the air-water interface, which often leads to detrimental effects such as denaturation or adoption of preferred orientations, ultimately hindering structure determination. Randomly biotinylating the protein of interest (for example, at its primary amines) and then tethering it to a cryo-EM grid coated with two-dimensional crystals of streptavidin (acting as an affinity surface) can prevent the protein from interacting with the air-water interface. Recently, this approach was successfully used to solve a high-resolution structure of a test sample, a bacterial ribosome. However, whether this method can be used for samples where interaction with the air-water interface has been shown to be problematic remains to be determined. Here we report a 3.1 Å structure of an RNA polymerase II elongation complex stalled at a cyclobutane pyrimidine dimer lesion (Pol II EC(CPD)) solved using streptavidin grids. Our previous attempt to solve this structure using conventional sample preparation methods resulted in a poor quality cryo-EM map due to Pol II EC(CPD)'s adopting a strong preferred orientation. Imaging the same sample on streptavidin grids improved the angular distribution of its view, resulting in a high-resolution structure. This work shows that streptavidin affinity grids can be used to address known challenges posed by the interaction with the air-water interface.