The solution structure of the pentatricopeptide repeat protein PPR10 upon binding atpH RNA

Abstract

The pentatricopeptide repeat (PPR) protein family is a large family of RNA-binding proteins that is characterized by tandem arrays of a degenerate 35-amino-acid motif which form an α-solenoid structure. PPR proteins influence the editing, splicing, translation and stability of specific RNAs in mitochondria and chloroplasts. Zea mays PPR10 is amongst the best studied PPR proteins, where sequence-specific binding to two RNA transcripts, atpH and psaJ, has been demonstrated to follow a recognition code where the identity of two amino acids per repeat determines the base-specificity. A recently solved ZmPPR10:psaJ complex crystal structure suggested a homodimeric complex with considerably fewer sequence-specific protein–RNA contacts than inferred previously. Here we describe the solution structure of the ZmPPR10:atpH complex using size-exclusion chromatography-coupled synchrotron small-angle X-ray scattering (SEC-SY-SAXS). Our results support prior evidence that PPR10 binds RNA as a monomer, and that it does so in a manner that is commensurate with a canonical and predictable RNA-binding mode across much of the RNA–protein interface.

An essential pentatricopeptide repeat protein in the apicomplexan remnant chloroplast

“…The protein eluted as a dimer from a gel filtration column (see below), and this dimerisation was confirmed by AUC following cleavage of the TRX‐His 6 tag by HRV 3C protease (Figure S6). The observed folding and dimerisation is consistent with other reported plant PPR proteins, though dimerisation may not occur in vivo (Barkan et al, ; Gully et al, ; Ke et al, ). A SWISS_MODEL homology modelling was carried out (Waterhouse et al, ) to compare both Pf PPR1 and Tg PPR1 with Arabidopsis thaliana PPR10, which has a solved structure.…”

Nuclear Protein-Only Ribonuclease P2 Structure and Biochemical Characterization Provide Insight into the Conserved Properties of tRNA 5′ End Processing Enzymes

“…The PPR domain of PRORP2 consists of five and a half PPR motifs corresponding to eleven consecutive α-helices similar to PRORP1. Based on the available structures of PPR motif containing proteins 8; 29; 30; 34; 35; 36 sequential PPR motifs packing in a parallel fashion results in a spiral of anti-parallel α-helices that form a right-handed superhelical structure. The two helices in a PPR motif are referred to as helix A and helix B, with all odd-numbered helices A in a superhelical PPR arrangement forming an inner concave surface while all even-numbered helices B are facing outwards and form the convex surface.…”

Spontaneous Dominant Mutations in Chlamydomonas Highlight Ongoing Evolution by Gene Diversification

“…While tetratricopeptide repeat (34 amino acids), Huntington, Elongation factor 3, protein phosphatase 2A, and yeast kinase TOR1 (HEAT; 39 amino acids), Armadillo (38 amino acids), Ankirin (33 amino acids), and leucine-rich repeat (23 to 24 amino acids) repeats are involved in protein-protein interactions, Pumilio and fem-3 binding factor (PUF; 36 amino acids), Transcription Activator Like Effector (TALE; 34 amino acids), pentatricopeptide repeat (PPR; 35 amino acids), Half A Tetratricopeptide (HAT; 34 amino acids), and mitochondrial termination factor (mTERF; ;30 amino acids) motifs mediate protein-nucleic acid interactions (reviewed in Rubinson and Eichman, 2012). Crystallographic structures of PUF (Wang et al, 2001(Wang et al, , 2002Miller et al, 2008), TALE (Deng et al, 2012;Mak et al, 2012), and PPR (Ke et al, 2013;Yin et al, 2013;Gully et al, 2015) proteins in complex with their RNA/DNA targets confirmed that nucleic acids bind in an extended conformation to the inner concave surface of the solenoid, with each nucleotide contacting one, or at most two, consecutive repeats. Thus, repeats act in a modular fashion, with each repeat interacting with one nucleotide.…”