*3.4. Prediction of Secondary Structure*

Jpred4 (http://www.compbio.dundee.ac.uk/jpred/) was used for prediction of secondary structure for each sequence in our dataset [43].

### *3.5. Analysis and Visualization*

Algorithms of search, collection, representation and analysis by the described methods of the data were realized using the freely available programming language Python 3 (https://www.python.org/). The result of the obtained two-dimensional array of data (for CH-CDF plots) was visualized using the Matplotlib library.

#### **4. Conclusions**

In this work, we show that S1 proteins belong to a unique family, which differs in the classical sense from proteins with tandem repeats. We found that the one-domain and two-domain containing S1 proteins apparently have more stable and rigid structure. An increase in the number of structural domains contributes to the possible transition of a portion of proteins from the folded state to the MG state. For example, for three- and four-domain containing proteins, the ratio of predicted MG state is about 70%. A relatively small percentage of internal flexibility/disorder within individual structural domains could be seen as an indicator of the stability of the S1 domain as one of the OB-fold in this family. At the same time the ratio of flexibility in the separate domains apparently is related to their roles in the activity and functionality of S1. A more stable, compact and conservative central part in the multi-domain proteins is vital for RNA interaction, while terminals domains are for other functions. At the same time, an equal ratio of regions connecting the secondary structure in separate domains and between structural domains indicates about the same organization of multi-domains containing S1 proteins, as well as position and ratio of the secondary structures within separate domains. Reasons for the lack of intact 3D structure of full-length ribosomal protein S1 is not well-understood Perhaps this is due to the high mobility of domains relative to each other in the multi-domain proteins. Further investigation of the flexibility of the available 3D structures for separate S1 domains and the full length S1 domain from *E. coli* in complex with 70S ribosomal subunit will allow finding an accurate explanation.

**Author Contributions:** Conceptualization and experiment design, O.G., A.M., E.D.; software, A.M., M.L.; formal analysis, A.M., M.L., E.D., O.G.; data analysis, A.M., E.D., O.G.; visualization, A.M.; writing—original draft preparation, E.D. and O.G.; writing—review & editing, O.G.; supervision, O.G.

**Funding:** This research was funded by the RUSSIAN SCIENCE FOUNDATION, grant number 18-14-00321.

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