N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Science, Moscow, Russia.
M.M. Shemyakin and Y.A. Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Science, Moscow, Russia.
Int J Biol Macromol. 2023 Jul 31;244:125403. doi: 10.1016/j.ijbiomac.2023.125403. Epub 2023 Jun 15.
The clinical isolate of Klebsiella pneumoniae 1333/P225 was revealed as containing a KL108 K. pneumoniae K locus for capsule biosynthesis. The gene cluster demonstrated a high level of sequence and arrangement similarity with that of the E. coli colanic acid biosynthesis gene cluster. The KL108 gene cluster includes a gene of WcaD polymerase responsible for joining oligosaccharide K units into capsular polysaccharide (CPS), acetyltransferase, pyruvyltransferasefive and genes for glycosyltransferases (Gtrs), four of which have homologues in genetic units of the colanic acid synthesis. The fifth Gtr is specific to this cluster. The work involved the use of sugar analysis, Smith degradation and one- and two-dimensional H and C NMR spectroscopy to establish the structure of the K108 CPS. The CPS repetitive K unit is composed of branched pentasaccharide with three monosaccharides in the backbone and a disaccharide side chain. The main chain is the same as for colanic acid but the side chain differs. Two bacteriophages infecting K. pneumoniae strain 1333/P225 were isolated and structural depolymerase genes were determined; depolymerases Dep108.1 and Dep108.2 were cloned, expressed and purified. It was demonstrated that both depolymerases specifically cleave the β-Glcp-(1→4)-α-Fucp linkage between K108 units in the CPS.
肺炎克雷伯菌 1333/P225 的临床分离株被揭示为含有 KL108 K. pneumoniae K 位点用于荚膜生物合成。该基因簇表现出与大肠杆菌 colanic 酸生物合成基因簇高度的序列和排列相似性。KL108 基因簇包括负责将寡糖 K 单位连接到荚膜多糖 (CPS) 的 WcaD 聚合酶基因、乙酰基转移酶、丙酮酸基转移酶和糖基转移酶基因 (Gtrs)，其中四个在 colanic 酸合成的遗传单元中具有同源物。第五个 Gtr 是该簇特有的。该工作涉及使用糖分析、Smith 降解和一维和二维 H 和 C NMR 光谱学来确定 K108 CPS 的结构。CPS 重复的 K 单位由具有三个单糖的支化五糖组成，主链与 colanic 酸相同，但侧链不同。两种感染肺炎克雷伯菌 1333/P225 的噬菌体被分离出来，并确定了结构解聚酶基因；解聚酶 Dep108.1 和 Dep108.2 被克隆、表达和纯化。证明这两种解聚酶都特异性地切割 CPS 中 K108 单位之间的β-Glcp-(1→4)-α-Fucp 键。
