Crystallographic characterization of CCG repeats

Abstract

CCG repeats are highly over-represented in exons of the human genome. Usually they are located in the 5′ UTR but are also abundant in translated sequences. The CCG repeats are associated with three tri-nucleotide repeat disorders: Huntington’s disease, myotonic dystrophy type 1 and chromosome X-linked mental retardation (FRAXE). In this study, we present two crystal structures containing double-stranded CCG repeats: one of an RNA in the native form, and one containing LNA nucleotides. Both duplexes form A-helices but with strands slipped in the 5′ (native structure) or the 3′ direction (LNA-containing structure). As a result, one of two expected C-C pairs is eliminated from the duplex. Each of the three observed C-C pairs interacts differently, forming either one weak H-bond or none. LNA nucleotides have no apparent effect on the helical parameters but the base stacking is increased compared to the native duplex and the distribution of electrostatic potential in the major groove is changed. The CCG crystal structures explain the thermodynamic fragility of CCG runs and throw light on the observation that the MBNL1 protein recognises CCG runs, as well as CUG and CAG, but not the relatively stable CGG repeats.

Watson–Crick-like pairs in CCUG repeats: evidence for tautomeric shifts or protonation

“…This is reminiscent of the two known crystal structures of CCG repeats: one containing an LNA residue and the other unmodified (Kiliszek et al 2012). In both structures strand slippage was observed, reducing the number of CCG repeats from two to one.…”

“…A bending of the CCUG helix was also reported by Childs-Disney et al (2014). The bending/coiling effect was not observed in any of the tri-nucleotide repeats analyzed before (Kiliszek et al 2009(Kiliszek et al , 2010(Kiliszek et al , 2011(Kiliszek et al , 2012.…”

An innate twist between Crick’s wobble and Watson-Crick base pairs

“…Such large values of Δt and Δr are expected to significantly affect RNA conformations. These might be relevant for protein recognition interactions, as nWC pairs like U·U, U·C, C·C, A·A are involved in trinucleotide repeat expansion diseases (Gatchel and Zoghbi 2005) such as myotonic dystrophy (Kumar et al 2011), fragile X syndrome E (Kiliszek et al 2012), Huntington's disease, dentatorubral-palidoluysian atrophy, and spinocerebellar ataxias (Kiliszek et al 2010). Besides Δt, one might also expect Δr to play a significant role on the global conformation of RNA duplexes comprising purinepurine or pyrimidine-pyrimidine pairs.…”

Advancing viral RNA structure prediction: measuring the thermodynamics of pyrimidine-rich internal loops

“…This type of pairing would not be possible if the N3 position were protonated, but possible C + C + pairs could form with two hydrogen bonds. There are several possible conformations for CC and CU pairs that would be disrupted by cytosine protonation (Kiliszek et al 2012;Rypniewski et al 2016). Cytidine protonation would also change stacking interactions.…”