Effect of different N7 substitution of dinucleotide cap analogs on the hydrolytic susceptibility towards scavenger decapping enzymes (DcpS)

Karolina Piecyk; Zbigniew M Darzynkiewicz; Marzena Jankowska-Anyszka; Aleksandra Ferenc-Mrozek; Janusz Stepinski; Edward Darzynkiewicz; Elzbieta Bojarska

doi:10.1016/j.bbrc.2015.06.001

Effect of different N7 substitution of dinucleotide cap analogs on the hydrolytic susceptibility towards scavenger decapping enzymes (DcpS)

Biochem Biophys Res Commun. 2015 Aug 14;464(1):89-93. doi: 10.1016/j.bbrc.2015.06.001. Epub 2015 Jun 4.

Authors

Karolina Piecyk¹, Zbigniew M Darzynkiewicz², Marzena Jankowska-Anyszka³, Aleksandra Ferenc-Mrozek⁴, Janusz Stepinski⁵, Edward Darzynkiewicz², Elzbieta Bojarska⁶

Affiliations

¹ Faculty of Chemistry, University of Warsaw, 1 Pasteura St., 02-093 Warsaw, Poland.
² Centre of New Technologies, University of Warsaw, 2c Banacha St., 02-097 Warsaw, Poland; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, 93 Zwirki & Wigury St., 02-089 Warsaw, Poland.
³ Faculty of Chemistry, University of Warsaw, 1 Pasteura St., 02-093 Warsaw, Poland; Department of Biochemistry, Second Faculty of Medicine, Medical University of Warsaw, 101 Zwirki & Wigury St., 02-089 Warsaw, Poland.
⁴ Centre of New Technologies, University of Warsaw, 2c Banacha St., 02-097 Warsaw, Poland; College of Inter-Faculty Individual Studies in Mathematics and Natural Sciences, University of Warsaw, 93 Zwirki & Wigury St., 02-089 Warsaw, Poland.
⁵ Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, 93 Zwirki & Wigury St., 02-089 Warsaw, Poland.
⁶ Centre of New Technologies, University of Warsaw, 2c Banacha St., 02-097 Warsaw, Poland. Electronic address: e.bojarska@cent.uw.edu.pl.

PMID: 26049109
DOI: 10.1016/j.bbrc.2015.06.001

Abstract

Scavenger decapping enzymes (DcpS) are involved in eukaryotic mRNA degradation process. They catalyze the cleavage of residual cap structure m(7)GpppN and/or short capped oligonucleotides resulting from exosom-mediated the 3' to 5' digestion. For the specific cap recognition and efficient degradation by DcpS, the positive charge at N7 position of guanine moiety is required. Here we examine the role the N7 substitution within the cap structure on the interactions with DcpS (human, Caenorhabditis elegans and Ascaris suum) comparing the hydrolysis rates of dinucleotide cap analogs (m(7)GpppG, et(7)GpppG, but(7)GpppG, bn(7)GpppG) and the binding affinities of hydrolysis products (m(7)GMP, et(7)GMP, but(7)GMP, bn(7)GMP). Our results show the conformational flexibility of the region within DcpS cap-binding pocket involved in the interaction with N7 substituted guanine, which enables accommodation of substrates with differently sized N7 substituents.

Keywords: Cap analogs; Decapping scavengers; Enzyme kinetic; Fluorescence titration; mRNA degradation.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Ascaris suum / genetics
Ascaris suum / metabolism
Caenorhabditis elegans / genetics
Caenorhabditis elegans / metabolism
Caenorhabditis elegans Proteins / chemistry*
Caenorhabditis elegans Proteins / genetics
Caenorhabditis elegans Proteins / metabolism
Endoribonucleases / chemistry*
Endoribonucleases / genetics
Endoribonucleases / metabolism
Enzyme Assays
Escherichia coli / genetics
Escherichia coli / metabolism
Gene Expression
Humans
Isoenzymes / chemistry
Isoenzymes / genetics
Isoenzymes / metabolism
Kinetics
Nucleic Acid Conformation
Pyrophosphatases / chemistry*
Pyrophosphatases / genetics
Pyrophosphatases / metabolism
RNA Cap Analogs / chemistry*
RNA Cap Analogs / metabolism
RNA Stability / genetics*
Recombinant Fusion Proteins / chemistry*
Recombinant Fusion Proteins / genetics
Recombinant Fusion Proteins / metabolism
Static Electricity

Substances

Caenorhabditis elegans Proteins
Isoenzymes
RNA Cap Analogs
Recombinant Fusion Proteins
Endoribonucleases
DcpS protein, human
Pyrophosphatases
dcs-1 protein, C elegans