Directed evolution of a dextransucrase for increased constitutive activity and the synthesis of a highly branched dextran

An Escherichia coli transformant (pDSRB742CK) was obtained from the DSRB742 clone by using ultrasoft X-rays for the expression of a dextransucrase. The enzyme differed in several aspects from DSRB742 dextransucrase: it (1) was constitutive; (2) was extracellular; (3) had 2.6 times greater activity (0.035 IU/ml and 0.23 IU/mg); and (4) synthesized a highly (15.6%) α-(1→3) branched dextran. Seven nucleotides of the parent gene (dsrB742) were changed in the nucleotide sequence; four nucleotides were changed in the open reading frame (ORF) that resulted in a 30 amino acid deletion in the N-terminus.