Engineered rings of mixed yeast Lsm proteins show differential interactions with translation factors and U-Rich RNA

Meghna Sobti, Liza Cubeddu, Paul A. Haynes, Bridget C. Mabbutt

Research output: Contribution to journalArticlepeer-review

10 Citations (Scopus)

Abstract

The Lsm proteins organize as heteroheptameric ring assemblies capable of binding RNA substrates and ancillary protein factors. We have constructed simplified Lsm polyproteins that organize as multimeric ring structures as analogues of the functional Lsm complexes. Polyproteins Lsm[2+3], Lsm[4+1], and Lsm[5+6] incorporate natural sequence extensions as linker peptides between the core Lsm domains. In solution, the recombinant products organize as stable ring oligomers (75 Å wide, 20 Å pores) in discrete tetrameric and octameric forms. Following immobilization, the polyproteins successfully act as affinity pulldown ligands for proteins within yeast lysáte, including native Lsm proteins. Interaction, partners were consistent with current models of the mixed Lsm ring assembly in vivo but also suggest that dynamic rearrangements of Lsm protein complexes can occur. The Lsm polyprotein ring complexes were seen, in gel shift assays to have a preference for U-rich RNA sequences, with, tightest binding measured for Lsm[2+3] with U10. Polyprotein rings containing truncated forms of Lsm1 and Lsm4 were found to associate with translation, initiation, and elongation protein factors in an RNA-dependent manner. Our findings suggest Lsm1 and/or Lsm4 can interact with translationally active mRNA.

Original languageEnglish
Pages (from-to)2335-2345
Number of pages11
JournalBiochemistry
Volume49
Issue number11
DOIs
Publication statusPublished - 23 Mar 2010

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