Biology subjects

Dienemann, C.

Publications and source records attributed to Dienemann, C..

21 records · Page 2Linked to original sources

bioRxiv2020

Structure of replicating SARS-CoV-2 polymerase

The coronavirus SARS-CoV-2 uses an RNA-dependent RNA polymerase (RdRp) for the replication of its genome and the transcription of its genes. Here we present the cryo-electron microscopic structure of the SARS-CoV-2 RdRp in its replicating form. The structure comprises the viral proteins nsp12, nsp8, and nsp7, and over two turns of RNA template-product duplex. The active site cleft of nsp12 binds the first turn of RNA and mediates RdRp activity with conserved residues. Two copies of nsp8 bind to opposite sides of the cleft and position the RNA duplex as it exits. Long helical extensions in nsp8 protrude along exiting RNA, forming positively charged sliding poles that may enable processive replication of the long coronavirus genome. Our results will allow for a detailed analysis of the inhibitory mechanisms used by antivirals such as remdesivir, which is currently in clinical trials for the treatment of coronavirus disease 2019 (COVID-19).

biophysics↗

bioRxiv2019

Structure of SWI/SNF chromatin remodeller RSC bound to a nucleosome

Chromatin remodelling complexes of the SWI/SNF family function in the formation of nucleosome-depleted regions and transcriptionally active promoters in the eukaryote genome. The structure of the Saccharomyces cerevisiae SWI/SNF family member RSC in complex with a nucleosome substrate reveals five protein modules and suggests key features of the remodelling mechanism. A DNA-interacting module grasps extra-nucleosomal DNA and helps to recruit RSC to promoters. The ATPase and arm modules sandwich the nucleosome disc with their SnAC and finger elements, respectively. The translocase motor engages with the edge of the nucleosome at superhelical location +2 to pump DNA along the nucleosome, resulting in a sliding of the histone octamer along DNA. The results elucidate how nucleosome-depleted regions are formed and provide a basis for understanding human chromatin remodelling complexes of the SWI/SNF family and the consequences of cancer mutations that frequently occur in these complexes.

molecular biology↗

bioRxiv2019

Structural basis of TFIIH activation for nucleotide excision repair

Genomes are constantly threatened by DNA damage, but cells can remove a large variety of DNA lesions by nucleotide excision repair (NER)1. Mutations in NER factors compromise cellular fitness and cause human diseases such as Xeroderma pigmentosum (XP), Cockayne syndrome and trichothiodystrophy2,3. The NER machinery is built around the multisubunit transcription factor IIH (TFIIH), which opens the DNA repair bubble, scans for the lesion, and coordinates excision of the damaged DNA single strand fragment1,4. TFIIH consists of a kinase module and a core module that contains the ATPases XPB and XPD5. Here we prepare recombinant human TFIIH and show that XPB and XPD are stimulated by the additional NER factors XPA and XPG, respectively. We then determine the cryo-electron microscopy structure of the human core TFIIH-XPA-DNA complex at 3.6 [A] resolution. The structure represents the lesion-scanning intermediate on the NER pathway and rationalizes the distinct phenotypes of disease mutations. It reveals that XPB and XPD bind double- and single-stranded DNA, respectively, consistent with their translocase and helicase activities. XPA forms a bridge between XPB and XPD, and retains the DNA at the 5-edge of the repair bubble. Biochemical data and comparisons with prior structures6,7 explain how XPA and XPG can switch TFIIH from a transcription factor to a DNA repair factor. During transcription, the kinase module inhibits the repair helicase XPD8. For DNA repair, XPA dramatically rearranges the core TFIIH structure, which reorients the ATPases, releases the kinase module and displaces a plug element from the DNA-binding pore in XPD. This enables XPD to move by ~80 [A], engage with DNA, and scan for the lesion in a XPG-stimulated manner. Our results provide the basis for a detailed mechanistic analysis of the NER mechanism.

molecular biology↗