Structure of Secondary Cell Wall Polymers in Cell Surfaces of Anaerobes and Interaction between Them and SLH Domains

1.从3株梭状芽胞杆菌、约氏梭状芽胞杆菌、空心杆菌和高温梭状芽胞杆菌中制备了次级细胞壁聚合物(SCWP)和肽多聚糖。2.从约氏梭菌木聚糖酶Xyn10A、杆菌木聚糖酶Xyn10B和高温梭状芽胞杆菌细胞表面蛋白SDBA出发，在大肠杆菌中制备了SLH模块多肽，并进行了纯化。结果发现，来自C.josui Xyn10A和C.stercorum Xyn10B的SLH模块多肽与SCWP结合，而来自致热梭菌SdbA的SLH模块多肽意外地与肽聚糖结合。4.利用从大肠杆菌中制备的多糖组分进行的结合分析证实，来自致热梭菌SdbA的SLH模块多肽与肽聚糖具有亲和力，尽管人们普遍认为SLH模块与SCWP结合。高温假单胞菌SdbA的SLH模块多肽可用于缺乏SCWP.5的细菌的细胞表面工程。用HCl对约氏乳杆菌、斯特氏假单胞菌和高温假单胞菌的SCWP进行了降解，并对其氨基酸组成进行了定性分析。检测到N-乙酰氨基葡萄糖和N-乙酰甘露糖胺是菌丝体SCWP和N-乙酰半乳糖胺的组成成分，而这些氨基糖在约氏梭菌和高温梭菌SCWP中得到了鉴定，表明这两种细菌的SCWP含有一个未知的结构。

1.Secondary cell wall polymers(SCWP) and peptidoglycans were prepared from 3 clostridia, C.josui, C.stercorarium and C.thermocellum2.SLH module polypeptides from C.josui xylanase Xyn10A, C.stercorarium xylanase Xyn10B and C.thermocellum cell surface protein SdbA were produced in Escherichia coli and purified.3.Interactions between these polypeptides and various cell wall fractions were investigated. As a result, it was found that the SLH module polypeptides from C.josui Xyn10A and C.stercorarium Xyn10B bound to SCWP but that the SLH module polypeptide from C.thermocellum SdbA unexpectedly bound to peptidoglycan.4.Binding assay using peptidoglycan fraction prepared from E.coli confirmed that the SLH module polypeptide from C.thermocellum SdbA had an affinity for peptidoglycan although it is generally believed that SLH modules bind to SCWP. The SLH module polypeptide from C.thermocellum SdbA should be applicable for cell surface engineering in bacteria lacking SCWP.5.SCWP of C.josui, C.stercorarium and C.thermocellum were hydrolyzed with HC1 and constituent amino sugars were qualitatively analyzed. N-Acetylglucosamine and N-acetylmannosamine were detected as constituents for C.stercorarium SCWP and N-acetylgalactosamine as well as these amino sugars were identified for C.josui and C.thermocellum SCWP, suggesting that SCWP of these two bacteria include an unidentified structure.