**5. Conclusions**

The C-terminal region of the cyanobacterial protein LrtA, a member of the HPF family, was a disordered domain that self-associated through a mechanism that involved some of its tyrosine residues. These findings clarify a number of important features of intact LrtA, including demonstrating, in an unambiguous way, the presence of two distinct and structurally different domains in this protein. Moreover, in the absence of studies of the N-terminal region (residues 1-101) our results suggest that the unstructured C-terminal one is the region driving the supramolecular self-organization of the whole protein. This study contributes to clarify the structural and stability features of proteins that interact and modulate the ribosome activity, and especially those belonging to the almost unexplored subfamily of long HPF proteins.

**Supplementary Materials:** Supplementary materials can be found at http://www.mdpi.com/1422-0067/19/12/ 3902/s1. There are four figures, showing: Figure S1: the far-UV CD spectra of C-LrtA at two concentrations; Figure S2: the size exclusion chromatograms of C-LrtA at different concentrations; Figure S3: details of the purification and self-association of C-LrtA (Figure S3), which include the SDS-PAGE gels of purified C-LrtB (Figure S3A); the BN-PAGE (Figure S3B); and the cross-linked protein (Figure S3C) at different reaction times; Figure S4: the ITC thermogram.

**Author Contributions:** Conceptualization, J.L.N. and B.R.; Methodology, J.L.N., A.M.G., F.H., A.A. and B.R.; Software, B.R.; Investigation, J.L.N., A.M.G., F.H., A.A. and B.R.; Writing-Original Draft Preparation, J.L.N., A.M.G., F.H., A.A. and B.R.; Writing-Review and Editing, J.L.N. and B.R.

**Funding:** This research was funded by Spanish Ministry of Economy and Competitiveness [CTQ2015-64445-R (to J.L.N.) and MAT2015-63704-P (to A.A.), with Fondo Social Europeo (ESF)], and by the Basque Government [IT-654-13 (to A.A.)].

**Acknowledgments:** B.R. acknowledges kind hospitality and use of computational resources in the European Magnetic Resonance Center (CERM), Sesto Fiorentino (Florence), Italy. J.L.N. thanks F. J. Florencio and M. I. Muro-Pastor for introducing to him the LrtA system. J.L.N. and F.H. thank Javier Gómez for helpful discussions. All the authors thank the three anonymous reviewers for their encouragement to formulate some hypotheses on the biological significance of the oligomerization in LrtA.

**Conflicts of Interest:** The authors declare no conflict of interest.

### **Abbreviations**

