A double mutantEst1, which is a plastic degrading cutinase-type esterase in Thermobifida alba, has been over-expressed in Escherichia coli. The recombinant protein was purified by a two-step protocol involving immobil...A double mutantEst1, which is a plastic degrading cutinase-type esterase in Thermobifida alba, has been over-expressed in Escherichia coli. The recombinant protein was purified by a two-step protocol involving immobilized metal affinity chromatography and cation-exchange chromatography, yielding 120 mg of protein per liter of bacterial culture. Crystals have been obtained by using the sitting-drop vapor-diffusion technique. Native diffraction data to 1.37 Åresolution were obtained at the BL44XU beam line of SPring-8 from a flash-frozen crystal at 100 K. The crystals belong to space group C2, with unit-cell parameters a = 127.2 Å, b = 42.1 Å, c = 63.2 Å, β = 114.7°, likely containing one Est1 double mutant (296 residues) per asymmetric unit.展开更多
The original online version of this article (Kitadokoro, K., Matsui, S., Osokoshi, R., Nakata, K. and Kamitani, S. (2018) Expression, Purification and Crystallization of Thermostable Mutant of Cutinase Est1 from Therm...The original online version of this article (Kitadokoro, K., Matsui, S., Osokoshi, R., Nakata, K. and Kamitani, S. (2018) Expression, Purification and Crystallization of Thermostable Mutant of Cutinase Est1 from Thermobifida alba. Advances in Bioscience and Biotechnology, 9, 215-223. https://doi.org/10.4236/abb.2018.95015) unfortunately contains some mistakes. The author wishes to correct the errors.展开更多
文摘A double mutantEst1, which is a plastic degrading cutinase-type esterase in Thermobifida alba, has been over-expressed in Escherichia coli. The recombinant protein was purified by a two-step protocol involving immobilized metal affinity chromatography and cation-exchange chromatography, yielding 120 mg of protein per liter of bacterial culture. Crystals have been obtained by using the sitting-drop vapor-diffusion technique. Native diffraction data to 1.37 Åresolution were obtained at the BL44XU beam line of SPring-8 from a flash-frozen crystal at 100 K. The crystals belong to space group C2, with unit-cell parameters a = 127.2 Å, b = 42.1 Å, c = 63.2 Å, β = 114.7°, likely containing one Est1 double mutant (296 residues) per asymmetric unit.
文摘The original online version of this article (Kitadokoro, K., Matsui, S., Osokoshi, R., Nakata, K. and Kamitani, S. (2018) Expression, Purification and Crystallization of Thermostable Mutant of Cutinase Est1 from Thermobifida alba. Advances in Bioscience and Biotechnology, 9, 215-223. https://doi.org/10.4236/abb.2018.95015) unfortunately contains some mistakes. The author wishes to correct the errors.