In vivo immobilization of D-hydantoinase in Escherichia coli(BIOCHEMICAL ENGINEERING)

概要

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D-p-Hydroxyphenylglycine (D-HPG) is a precursor required for the synthesis of semi-synthetic antibiotics. This unnatural amino acid can be produced by a transformation reaction mediated by D-hydantoinase (D-HDT) and D-amidohydrolase. In this study, a method was developed to integrate production and immobilization of recombinant D-HDT in vivo. This was approached by first fusion of the gene encoding D-HDT with phaP (encoding phasin) of Ralstonia eutropha H16. The fusion gene was then expressed in the Escherichia coli strain that carried a heterologous synthetic pathway for polyhydroxyalkanoate (PHA). As a result, D-HDT was found to associate with isolated PHA granules. Further characterization illustrated that D-HDT immobilized on PHA exhibited the maximum activity at pH 9 and 60℃ and had a half-life of 95 h at 40℃. Moreover, PHA-bound D-HDT could be reused for 8 times with the conversion yield exceeding 90%. Overall, it illustrates the feasibility of this approach to facilitate in vivo immobilization of enzymes in heterologous E. coli strain, which may open a new avenue of enzyme application in industry.
公益社団法人日本生物工学会の論文