5osy
Human Decapping Scavenger enzyme (hDcpS) in complex with m7G(5'S)ppSp(5'S)G mRNA 5' cap analog
Structural highlights
FunctionDCPS_HUMAN Decapping scavenger enzyme that catalyzes the cleavage of a residual cap structure following the degradation of mRNAs by the 3'->5' exosome-mediated mRNA decay pathway. Hydrolyzes cap analog structures like 7-methylguanosine nucleoside triphosphate (m7GpppG) with up to 10 nucleotide substrates (small capped oligoribonucleotides) and specifically releases 5'-phosphorylated RNA fragments and 7-methylguanosine monophosphate (m7GMP). Cleaves cap analog structures like tri-methyl guanosine nucleoside triphosphate (m3(2,2,7)GpppG) with very poor efficiency. Does not hydrolyze unmethylated cap analog (GpppG) and shows no decapping activity on intact m7GpppG-capped mRNA molecules longer than 25 nucleotides. Does not hydrolyze 7-methylguanosine diphosphate (m7GDP) to m7GMP (PubMed:22985415). May also play a role in the 5'->3 mRNA decay pathway; m7GDP, the downstream product released by the 5'->3' mRNA mediated decapping activity, may be also converted by DCPS to m7GMP (PubMed:14523240). Binds to m7GpppG and strongly to m7GDP. Plays a role in first intron splicing of pre-mRNAs. Inhibits activation-induced cell death.[1] [2] [3] [4] [5] [6] [7] [8] [9] [10] Publication Abstract from PubMedThe 5' cap consists of 7-methylguanosine (m(7)G) linked by a 5'-5'-triphosphate bridge to messenger RNA (mRNA) and acts as the master regulator of mRNA turnover and translation initiation in eukaryotes. Cap analogues that influence mRNA translation and turnover (either as small molecules or as part of an RNA transcript) are valuable tools for studying gene expression, which is often also of therapeutic relevance. Here, we synthesized a series of 15 dinucleotide cap (m(7)GpppG) analogues containing a 5'-phosphorothiolate (5'-PSL) moiety (i.e., an O-to-S substitution within the 5'-phosphoester) and studied their biological properties in the context of three major cap-binding proteins: translation initiation factor 4E (eIF4E) and two decapping enzymes, DcpS and Dcp2. While the 5'-PSL moiety was neutral or slightly stabilizing for cap interactions with eIF4E, it significantly influenced susceptibility to decapping. Replacing the gamma-phosphoester with the 5'-PSL moiety (gamma-PSL) prevented beta-gamma-pyrophosphate bond cleavage by DcpS and conferred strong inhibitory properties. Combining the gamma-PSL moiety with alpha-PSL and beta-phosphorothioate (PS) moiety afforded first cap-derived hDcpS inhibitor with low nanomolar potency. Susceptibility to Dcp2 and translational properties were studied after incorporation of the new analogues into mRNA transcripts by RNA polymerase. Transcripts containing the gamma-PSL moiety were resistant to cleavage by Dcp2. Surprisingly, superior translational properties were observed for mRNAs containing the alpha-PSL moiety, which were Dcp2-susceptible. The overall protein expression measured in HeLa cells for this mRNA was comparable to mRNA capped with the translation augmenting beta-PS analogue reported previously. Overall, our study highlights 5'-PSL as a synthetically accessible cap modification, which, depending on the substitution site, can either reduce susceptibility to decapping or confer superior translational properties on the mRNA. The 5'-PSL-analogues may find application as reagents for the preparation of efficiently expressed mRNA or for investigation of the role of decapping enzymes in mRNA processing or neuromuscular disorders associated with decapping. 5'-Phosphorothiolate Dinucleotide Cap Analogues: Reagents for Messenger RNA Modification and Potent Small-Molecular Inhibitors of Decapping Enzymes.,Wojtczak BA, Sikorski PJ, Fac-Dabrowska K, Nowicka A, Warminski M, Kubacka D, Nowak E, Nowotny M, Kowalska J, Jemielity J J Am Chem Soc. 2018 May 1. doi: 10.1021/jacs.8b02597. PMID:29676910[11] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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