ABSTRACT
The rice xylanase inhibitor gene, rixi, was cloned from rice genome. The open reading frame of rixi was 915â¯bp and encoded 304 amino acids with the theoretical molecular mass of 33.9â¯kDa. The rixi was inserted into the new-type expression vector pCold TF, and was high-level expressed in Escherichia coli BL21 (DE3). SDS-PAGE and Western blot analysis revealed that the molecular weight of the recombinant rice xylanase inhibitor, namely reERIXI, was approximately 89.8â¯kDa. The reERIXI exhibited significant inhibitory activities against several family GH11 xylanases. After interaction with reERIXI, the residual activity of reBaxA50 and TfxA_CD214 were 59.24% and 44.41%, respectively. The optimal temperature of reERIXI inhibitory activity to reBaxA50 and TfxA_CD214 were 60⯰C and 50⯰C, respectively. The thermostability assay revealed that reERIXI was stable below 60⯰C. reERIXI showed high inhibitory when interacting with reBaxA50 and TfxA_CD214 for 30-60â¯min. The intrinsic fluorescence spectroscopy of reBaxA50 and TfxA_CD214 was quenched with increasing reERIXI concentration. Circular dichroism measurement revealed that ratio of helix of reBaxA50 and ratio of beta of TfxA_CD214 significantly decreased when interacting with reERIXI. The total concentration of hydrolytic products from beechwood xylan decreased when reERIXI was added.