The β1 domain of protein G can replace the chorismate mutase domain of the T-protein

T-protein is composed of chorismate mutase (AroQT) fused to the N-terminus of prephenate dehydrogenase (TyrA). Here, we report the replacement of AroQT with the β1-domain of protein G (Gβ1). The TyrA domain shows a strong dehydrogenase activity within the context of this fusion, and our data indicate that Gβ1-TyrA folds into a dimeric conformation. Amino acid substitutions in the Gβ1 domain of Gβ1-TyrA identified residues involved in stabilizing the TyrA dimeric conformation. Gβ1 substitutions in the N-terminal β-hairpin eliminated Gβ1-TyrA expression, whereas Gβ1-TyrA tolerated Gβ1 substitutions in the C-terminal β-hairpin and in the α-helix. All of the characterized variants folded into a dimeric conformation. The importance of the β2-strand in forming a Gβ1 homo-dimerization interface explains the relevance of the first-β-hairpin in stabilizing the dimeric TyrA protein.

► Gβ1 can replace AroQT in the T-protein. ► AroQT/TyrA specific-contact interface is not important to maintain a functional T-protein. ► Gβ1 N-terminal β-hairpin residues are important to maintain Gβ1-TyrA expression. ► Gβ1-TyrA expression tolerates substitutions at the C-terminal β-hairpin of the Gβ1 domain. ► The different Gβ1-TyrA protein variants fold in a dimeric conformation.