Abstract:In the present study, we utilized a previously characterized N-acetylglucosaminidase (AcmA) to develop a whole-cell catalyst of bacterial superoxide dismutase (SOD) in Lactococcus lactis. The truncated C-terminal domain (cA) and the full-length AcmA from L. lactis wild-type strain MB191, were used as the anchoring motifs to immobilize an Escherichia coli-derived SOD onto the surfaces of L. lactis ATCC11454 cells. The PCR-amplified cA fragment, the signal and the full-length sequences of acmA, were fused with sod to generate the recombinant ss-cA-sod and acmA-sod, respectively, followed by ligating into the expression vector pMG36K, yielding the recombinant strain MB193 (harboring the ss-cA-sod fusion gene) and MB194 (harboring acmA-sod), respectively. SDS-PAGE analysis showed that the substantial expression profile of the fusion enzymes cA-SOD and AcmA-SOD in the recombinant L. lactis MB193 and MB194, with the predicted Mr of 46 kD and 64 kD, respectively. The Mn-SOD activities of MB193 and MB194 cells were determined by using the standard xanthinoxidase assay procedure. It showed that MB193 and MB194 exhibited the higher whole-cell Mn-SOD activities [by (2.63 ± 0.51) U/mL and (3.51 ± 0.64) U/mL, respectively, compared to the control strain MB192 by (1.53 ± 0.38) U/mL] that expressing the intracellular SOD. In addition, the recombinant MB194 cells exhibited the higher cell-surface display efficiency (by 56.4%) compared to MB193 cells (by 41.9%).
