Solution structure of human P1•P2 heterodimer provides insights into the role of eukaryotic stalk in recruiting the ribosome-inactivating protein trichosanthin to the ribosome

Lateral ribosomal stalk is responsible for binding and recruiting translation factors during protein synthesis. The eukaryotic stalk consists of one P0 protein with two copies of P1 center dot P2 heterodimers to form a P0(P1 center dot P2)(2) pentameric P-complex. Here, we have solved the structure of full-length P1 center dot P2 by nuclear magnetic resonance spectroscopy. P1 and P2 dimerize via their helical N-terminal domains, whereas the C-terminal tails of P1 center dot P2 are unstructured and can extend up to similar to 125 A away from the dimerization domains. N-15 relaxation study reveals that the C-terminal tails are flexible, having a much faster internal mobility than the N-terminal domains. Replacement of prokaryotic L10(L7/L12)(4)/L11 by eukaryotic P0(P1 center dot P2)(2)/eL12 rendered Escherichia coli ribosome, which is insensitive to trichosanthin (TCS), susceptible to depurination by TCS and the C-terminal tail was found to be responsible for this depurination. Truncation and insertion studies showed that depurination of hybrid ribosome is dependent on the length of the proline-alanine rich hinge region within the C-terminal tail. All together, we propose a model that recruitment of TCS to the sarcin-ricin loop required the flexible C-terminal tail, and the proline-alanine rich hinge region lengthens this C-terminal tail, allowing the tail to sweep around the ribosome to recruit TCS.