Structural characterization of human ribosomal protein P2 by NMR spectroscopy

Abstract

Ribosome consists of rRNAs and ribosomal proteins. Among the ribosomal proteins, three acidic ribosomal proteins P0, P1 and P2, which form P-complex, are located at the lateral stalk of the ribosome. The P-complex consists of one P0, two P1 and two P2, but the protein-protein interactions among these five proteins are still controversial. On the functional point of view, it has been reported that these proteins interact with mRNAs, tRNAs and translation factors during the course of protein synthesis. To understand the structure-function relationship, P2 was overexpressed in E. coli and purified by four chromatographic steps including ion-exchange, hydrophobic interaction and gel filtration chromatography. P2 was labeled with C13 and/or N15 isotopes and NMR experiments were performed. By triple resonance experiments, we have obtained the backbone 1H, 13C, and 15N resonances of P2. Secondary chemical shift values suggested that P2 has an N-terminal domain with four helices, and an unstructured C-terminal tail. Pull-down assay suggested that the N-terminal domain of P2 is responsible for dimerization with P1 to form a heterodimer, or with P2 to form a homodimer. The C-terminal tail of P2 was found to interact with eukaryotic elongation factor 2 (eEF2), which is an important factor in protein elongation.