A second hybrid-binding domain modulates the activity of Drosophila ribonuclease H1

Show full item record



Permalink

http://hdl.handle.net/10138/323992

Citation

Gonzalez de Cozar , J M , Carretero-Junquera , M , Ciesielski , G L , Miettinen , S M , Varjosalo , M , Kaguni , L S , Dufour , E & Jacobs , H T 2020 , ' A second hybrid-binding domain modulates the activity of Drosophila ribonuclease H1 ' , Journal of Biochemistry , vol. 168 , no. 5 , pp. 515-533 . https://doi.org/10.1093/jb/mvaa067

Title: A second hybrid-binding domain modulates the activity of Drosophila ribonuclease H1
Author: Gonzalez de Cozar, Jose M.; Carretero-Junquera, Maria; Ciesielski, Grzegorz L.; Miettinen, Sini M.; Varjosalo, Markku; Kaguni, Laurie S.; Dufour, Eric; Jacobs, Howard T.
Contributor: University of Helsinki, Molecular Systems Biology
University of Helsinki, Biosciences
Date: 2020-11
Language: eng
Number of pages: 19
Belongs to series: Journal of Biochemistry
ISSN: 0021-924X
URI: http://hdl.handle.net/10138/323992
Abstract: In eukaryotes, ribonuclease H1 (RNase H1) is involved in the processing and removal of RNA/DNA hybrids in both nuclear and mitochondrial DNA. The enzyme comprises a C-terminal catalytic domain and an N-terminal hybrid-binding domain (HBD), separated by a linker of variable length, 115 amino acids in Drosophila melanogaster (Dm). Molecular modelling predicted this extended linker to fold into a structure similar to the conserved HBD. Based on a deletion series, both the catalytic domain and the conserved HBD were required for high-affinity binding to heteroduplex substrates, while loss of the novel HBD led to an similar to 90% drop in K-cat with a decreased K-M, and a large increase in the stability of the RNA/DNA hybrid-enzyme complex, supporting a bipartite-binding model in which the second HBD facilitates processivity. Shotgun proteomics following in vivo cross-linking identified single-stranded DNA-binding proteins from both nuclear and mitochondrial compartments, respectively RpA-70 and mtSSB, as prominent interaction partners of Dm RNase H1. However, we were not able to document direct and stable interactions with mtSSB when the proteins were cooverexpressed in S2 cells, and functional interactions between them in vitro were minor.
Subject: biolayer interferometry
mitochondria
ribonuclease H
shotgun proteomics
single-stranded DNA-binding protein
HUMAN RNASE H1
MITOCHONDRIAL-DNA REPLICATION
MURINE LEUKEMIA-VIRUS
DOUBLE-STRANDED-RNA
MTDNA REPLICATION
POLYMERASE-GAMMA
RNA/DNA HYBRID
R-LOOPS
CRYSTAL-STRUCTURE
PROTEIN-A
1182 Biochemistry, cell and molecular biology
Rights:


Files in this item

Total number of downloads: Loading...

Files Size Format View
mvaa067.pdf 1.464Mb PDF View/Open

This item appears in the following Collection(s)

Show full item record