Effect of the disease-causing mutations identified in human ribonuclease (RNase) H2 on the activities and stabilities of yeast RNase H2 and archaeal RNase HII.

The FEBS Journal
Muhammad S RohmanShigenori Kanaya

Abstract

Eukaryotic ribonuclease (RNase) H2 consists of one catalytic and two accessory subunits. Several single mutations in any one of these subunits of human RNase H2 cause Aicardi-Goutières syndrome. To examine whether these mutations affect the complex stability and activity of RNase H2, three mutant proteins of His-tagged Saccharomyces cerevisiae RNase H2 (Sc-RNase H2*) were constructed. Sc-G42S*, Sc-L52R*, and Sc-K46W* contain single mutations in Sc-Rnh2Ap*, Sc-Rnh2Bp*, and Sc-Rnh2Cp*, respectively. The genes encoding the three subunits were coexpressed in Escherichia coli, and Sc-RNase H2* and its derivatives were purified in a heterotrimeric form. All of these mutant proteins exhibited enzymatic activity. However, only the enzymatic activity of Sc-G42S* was greatly reduced compared to that of the wild-type protein. Gly42 is conserved as Gly10 in Thermococcus kodakareansis RNase HII. To analyze the role of this residue, four mutant proteins, Tk-G10S, Tk-G10A, Tk-G10L, and Tk-G10P, were constructed. All mutant proteins were less stable than the wild-type protein by 2.9-7.6 degrees C in T(m). A comparison of their enzymatic activities, substrate binding affinities, and CD spectra suggests that the introduction of a bulky side chai...Continue Reading

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Citations

Nov 27, 2008·Journal of Molecular Medicine : Official Organ of the Gesellschaft Deutscher Naturforscher Und Ärzte·Fred W PerrinoThomas Hollis
Dec 24, 2010·The Journal of Biological Chemistry·Martin A M ReijnsAndrew P Jackson
Jan 29, 2013·Nucleic Acids Research·Hyongi ChonSusana M Cerritelli
Jul 10, 2010·BMC Evolutionary Biology·Jun OkadaShigenori Kanaya
Feb 18, 2016·The Journal of Experimental Medicine·Vladislav PokatayevRobert J Crouch
Oct 20, 2012·DNA Repair·Mercedes E AranaThomas A Kunkel
Nov 26, 2015·Progress in Biophysics and Molecular Biology·Shaolong Feng, Zhaohui Cao
Feb 21, 2009·The FEBS Journal·Susana M Cerritelli, Robert J Crouch
Aug 12, 2014·Biochemical Society Transactions·Martin A M Reijns, Andrew P Jackson
Dec 15, 2017·Developmental Neurobiology·Nicole A SuarezAlysson R Muotri
Jun 13, 2012·The FEBS Journal·Nujarin JongrujaShigenori Kanaya
Apr 2, 2011·The Journal of Biological Chemistry·Stephanie R CoffinFred W Perrino
Aug 3, 2019·DNA Repair·Malwina HyjekMarcin Nowotny

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