Substrate adaptabilities of Thermotogae mannan binding proteins as a function of their evolutionary histories

The Thermotogae possess a large number of ATP-binding cassette (ABC) transporters, including two mannan binding proteins, ManD and CelE (previously called ManE). We show that a gene encoding an ancestor of these was acquired by the Thermotogae from the archaea followed by gene duplication. To address the functional evolution of these proteins as a consequence of their evolutionary histories, we measured the binding affinities of ManD and CelE orthologs from representative Thermotogae. Both proteins bind cellobiose, cellotriose, cellotetraose, β-1,4-mannotriose, and β-1,4-mannotetraose. The CelE orthologs additionally bind β-1,4-mannobiose, laminaribiose, laminaritriose and sophorose while the ManD orthologs additionally only weakly bind β-1,4-mannobiose. The CelE orthologs have higher unfolding temperatures than the ManD orthologs. An examination of codon sites under positive selection revealed that many of these encode residues located near or in the binding site, suggesting that the proteins experienced selective pressures in regions that might have changed their functions. The gene arrangement, phylogeny, binding properties, and putative regulatory networks suggest that the ancestral mannan binding protein was a CelE ortholog which gave rise to the ManD orthologs. This study provides a window on how one class of proteins adapted to new functions and temperatures to fit the physiologies of their new hosts.