Intradiol dioxygenases (EC 1.13.11.1) are bacterial enzymes that catalyze the ring cleavage of catechols which is a central step in the aerobic degradation of aromatic compounds. Some members of this enzyme group have a C-terminus which is 4-5% longer (an additional 13-18 amino acids) compared to the majority of known sequences. The longer C-terminus itself is not highly conserved and appears to be poorly integrated in the protein structural models developed for representative intradiol dioxygenases. Using a protein engineering approach variant intradiol dioxygenases were produced by truncating the C-terminus to a size comparable to the shorter versions of the enzyme. Three enzymes were selected and were origin... More
Intradiol dioxygenases (EC 1.13.11.1) are bacterial enzymes that catalyze the ring cleavage of catechols which is a central step in the aerobic degradation of aromatic compounds. Some members of this enzyme group have a C-terminus which is 4-5% longer (an additional 13-18 amino acids) compared to the majority of known sequences. The longer C-terminus itself is not highly conserved and appears to be poorly integrated in the protein structural models developed for representative intradiol dioxygenases. Using a protein engineering approach variant intradiol dioxygenases were produced by truncating the C-terminus to a size comparable to the shorter versions of the enzyme. Three enzymes were selected and were originally described from the model organisms; Burkholderia xenovorans LB400， Pseudomonas putida KT2440 and Acinetobacter baylyi ADP1. The activity of the truncated enzymes were compared to the unmodified enzymes which revealed that truncation of the C-terminus could alter the enzyme activity; increasing the LB400 enzyme activity by as much as five fold， but reducing the activity of the intradiol dioxygenases from KT2440 and ADP1. The difference in effect is explained by the presence of a greater number of amino acid residues that can contribute to forming stable protein structures in the KT2440 and ADP1 enzymes. It is hypothesized that C-terminal truncation could in some cases provide a useful strategy for increasing intradiol dioxygenase activity for biotechnological production of muconic and adipic acids.
