Advanced and easy-to-use web-server for bioinformatic analysis of subfamily-specific
and conserved positions in diverse protein superfamilies


This web-server is free and open to all users with no login requirement

... to identify and prioritize subfamily-specific and conserved positions in a functionally diverse superfamily
and to select hot-spots for rational design of the query protein

Version 2.0 since November 20th, 2019

Publication: D. Suplatov, Y. Sharapova, E. Geraseva, V. Švedas (2020) Zebra2: advanced and easy-to-use web-server for bioinformatic analysis of subfamily-specific and conserved positions in diverse protein superfamilies. Nucleic Acids Res., 48 (W1), W65–W71. DOI: 10.1093/nar/gkaa276



  • The Zebra2 (i.e., Zebra v.2) is a highly automated web-tool to search for subfamily-specific and conserved positions (i.e., the determinants of functional diversity as well as the key catalytic and structural residues) in protein superfamilies;
  • Information about both conserved and specific positions can help to understand how the enzyme performs its natural function, while the latter can also be selected as hotspots for directed evolution or rational design experiments in an attempt to improve the wild-type protein variant for a particular purpose;
  • The bioinformatic analysis is supported by the integrated Mustguseal web-server to construct large structure-guided sequence alignments of functionally diverse families that can include thousands of proteins based on all available information in public databases;
  • The Zebra2 results are automatically prioritized by the bioinformatic analysis and provided at four information levels to facilitate the knowledge-driven expert selection of the most promising positions on-line: as a sequence similarity network; interfaces to sequence-based and 3D-structure-based analysis of conservation and variability; and accompanied by the detailed annotation of proteins accumulated from the integrated databases with links to the external resources (i.e., PDB, Uniprot, BacDive, BRENDA);
  • The integration of web-based bioinformatic tools Zebra2 and Mustguseal provides the first out-of-the-box open-access solution to identify subfamily-specific and conserved positions in protein superfamilies – to study function-associated patterns of local structure, to assist at selecting hot-spots for rational design and to prepare focused libraries for directed evolution.


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