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Chitoporin from Serratia marcescens: recombinant expression, purification and crystallization
Acta Crystallographica Section F ( IF 1.1 ) Pub Date : 2020-11-02 , DOI: 10.1107/s2053230x20013874 Rawiporn Amornloetwattana 1 , Robert C Robinson 1 , Hannadige Sasimali Madusanka Soysa 2 , Bert van den Berg 3 , Wipa Suginta 1
Acta Crystallographica Section F ( IF 1.1 ) Pub Date : 2020-11-02 , DOI: 10.1107/s2053230x20013874 Rawiporn Amornloetwattana 1 , Robert C Robinson 1 , Hannadige Sasimali Madusanka Soysa 2 , Bert van den Berg 3 , Wipa Suginta 1
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Serratia marcescens is an opportunistic pathogen that commonly causes hospital‐acquired infections and can utilize chitin‐enriched nutrients as an alternative energy source. This study reports the identification of a chitoporin (ChiP), termed SmChiP, from the outer membrane of S. marcescens. Sequence alignment with genetically characterized ChiPs suggests that SmChiP is more closely related to the monomeric EcChiP from Escherichia coli than to the trimeric VhChiP from Vibrio campbellii. A single crystal of SmChiP grown under the condition 22%(w/v) PEG 8000, 0.1 M calcium acetate, 0.1 M MES pH 6.0 diffracted X‐ray synchrotron radiation to 1.85 Å resolution. SmChiP co‐crystallized with chitohexaose under the condition 19%(w/v) PEG 1500, 2 M ammonium phosphate monobasic, 0.1 M HEPES pH 7.0 diffracted X‐rays to 2.70 Å resolution. Preliminary crystallographic analysis shows that both SmChiP crystal forms contain one molecule per asymmetric unit and that they belong to the tetragonal space groups P42212 and P41212, respectively. The SmChiP crystal has unit‐cell parameters a = 82.97, b = 82.97, c = 189.53 Å, α = β = γ = 90°, while the crystal of SmChiP in complex with chitohexaose has unit‐cell parameters a = 73.24, b = 73.24, c = 213.46 Å, α = β = γ = 90°. Initial assessment of the complex structure clearly revealed electron density for the sugar ligand. Structure determination of SmChiP in the absence and presence of chitohexaose should reveal the molecular basis of chitin utilization by S. marcescens.
中文翻译：
来自粘质沙雷氏菌的壳孔素：重组表达、纯化和结晶
粘质沙雷氏菌是一种机会性病原体，通常会引起医院获得性感染，可以利用富含几丁质的营养物质作为替代能源。本研究报告了从粘质沙门氏菌外膜中鉴定出的壳孔素 (ChiP)，称为Sm ChiP 。与遗传特征 ChiP 的序列比对表明，Sm ChiP 与大肠杆菌的单体Ec ChiP 的相关性比与坎贝氏弧菌的三聚体Vh ChiP 的相关性更密切。在 22%（ w / v）PEG 8000、0.1 M醋酸钙、0.1 M MES pH 6.0条件下生长的Sm ChiP单晶，衍射 X 射线同步加速器辐射分辨率为 1.85 Å。Sm ChiP 与壳六糖在 19%( w / v ) PEG 1500、2 M磷酸二氢铵、0.1 M HEPES pH 7.0条件下共结晶，衍射 X 射线分辨率为 2.70 Å。初步晶体学分析表明，两种Sm ChiP晶型每个不对称单元均含有一个分子，并且它们分别属于四方空间群P 4 2 2 1 2和P 4 1 2 1 2。Sm ChiP晶体的晶胞参数a = 82.97，b = 82.97，c = 189.53 Å，α = β = γ = 90°，而与壳六糖复合的Sm ChiP晶体的晶胞参数a = 73.24，b = 73.24，c = 213.46 Å，α = β = γ = 90°。对复杂结构的初步评估清楚地揭示了糖配体的电子密度。在不存在和存在壳六糖的情况下， Sm ChiP的结构测定应该揭示粘质沙门氏菌利用几丁质的分子基础。
更新日期：2020-11-02
中文翻译：
来自粘质沙雷氏菌的壳孔素：重组表达、纯化和结晶
粘质沙雷氏菌是一种机会性病原体，通常会引起医院获得性感染，可以利用富含几丁质的营养物质作为替代能源。本研究报告了从粘质沙门氏菌外膜中鉴定出的壳孔素 (ChiP)，称为Sm ChiP 。与遗传特征 ChiP 的序列比对表明，Sm ChiP 与大肠杆菌的单体Ec ChiP 的相关性比与坎贝氏弧菌的三聚体Vh ChiP 的相关性更密切。在 22%（ w / v）PEG 8000、0.1 M醋酸钙、0.1 M MES pH 6.0条件下生长的Sm ChiP单晶，衍射 X 射线同步加速器辐射分辨率为 1.85 Å。Sm ChiP 与壳六糖在 19%( w / v ) PEG 1500、2 M磷酸二氢铵、0.1 M HEPES pH 7.0条件下共结晶，衍射 X 射线分辨率为 2.70 Å。初步晶体学分析表明，两种Sm ChiP晶型每个不对称单元均含有一个分子，并且它们分别属于四方空间群P 4 2 2 1 2和P 4 1 2 1 2。Sm ChiP晶体的晶胞参数a = 82.97，b = 82.97，c = 189.53 Å，α = β = γ = 90°，而与壳六糖复合的Sm ChiP晶体的晶胞参数a = 73.24，b = 73.24，c = 213.46 Å，α = β = γ = 90°。对复杂结构的初步评估清楚地揭示了糖配体的电子密度。在不存在和存在壳六糖的情况下， Sm ChiP的结构测定应该揭示粘质沙门氏菌利用几丁质的分子基础。
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