Irreversible substrate engineering

In addition to protein engineering, the substrate mimicking approach was also appHed for RhuA catalyst It was uncovered that RhuA can perform the aldol addition of DHA to aldehyde at remarkably high rates when the reactions were carried out in the presence of borate [29]. Indeed, when sodium borate was added, the rates of aldol formation improved between 35- and 100-fold [25]. Besides the intrinsic tolerance of RhuA for DHA, the measured retroaldol rates for some aldol adducts in the presence of borate were low or negligible as compared with the synthetic ones, making the process virtually irreversible [29, 30]. Therefore, it was further suggested that the aldol adduct may be trapped by the formation of borate complexes which would be less active substrates for the aldolase [29, 30]. 

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