Department of Chemistry, Texas A&M University, College Station, Texas 77843, United States.
Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 553706, United States.
Biochemistry. 2021 May 18;60(19):1552-1563. doi: 10.1021/acs.biochem.1c00183. Epub 2021 Apr 26.
is the leading cause of food poisoning in the United States and Europe. The exterior cell surface of is coated with a capsular polysaccharide (CPS) that is essential for the maintenance and integrity of the bacterial cell wall and evasion of the host immune response. The identity and sequences of the monosaccharide components of the CPS are quite variable and dependent on the specific strain of . It is currently thought that the immediate precursor for the multiple variations found in the heptose moieties of the CPS is GDP-d--α-d--heptose. In NCTC 11168, the heptose moiety is d--l--heptose. It has previously been shown that Cj1427 catalyzes the oxidation of GDP-d--α-d--heptose to GDP-d--4-keto-α-d--heptose using α-ketoglutarate as a cosubstrate. Cj1430 was now demonstrated to catalyze the double epimerization of this product at C3 and C5 to form GDP-d--4-keto-β-l--heptose. Cj1428 subsequently catalyzes the stereospecific reduction of this GDP-linked heptose by NADPH to form GDP-d--β-l--heptose. The three-dimensional crystal structure of Cj1430 was determined to a resolution of 1.85 Å in the presence of bound GDP-d--β-l--heptose, a product analogue. The structure shows that it belongs to the cupin superfamily. The three-dimensional crystal structure of Cj1428 was solved in the presence of NADPH to a resolution of 1.50 Å. Its fold places it into the short-chain dehydrogenase/reductase superfamily. Typically, members in this family display a characteristic signature sequence of YXXXK, with the conserved tyrosine serving a key role in catalysis. In Cj1428, this residue is a phenylalanine.
是美国和欧洲食源性疾病的主要致病菌。 的外表面覆盖有荚膜多糖（CPS），这对于维持和完整的细菌细胞壁和逃避宿主免疫反应是必不可少的。CPS 的单糖成分的身份和序列变化很大，取决于 的特定菌株。目前认为，在 CPS 的庚糖部分中发现的多种变化的直接前体是 GDP-d--α-d--庚糖。在 NCTC 11168 中，庚糖部分是 d--l--庚糖。先前已经表明，Cj1427 使用 α-酮戊二酸作为共底物，催化 GDP-d--α-d--庚糖氧化为 GDP-d--4-酮-α-d--庚糖。现在表明 Cj1430 催化该产物在 C3 和 C5 处的双重差向异构化为 GDP-d--4-酮-β-l--庚糖。Cj1428 随后催化 NADPH 对该 GDP 连接的庚糖进行立体特异性还原，形成 GDP-d--β-l--庚糖。在存在结合的 GDP-d--β-l--庚糖的情况下，测定了 Cj1430 的三维晶体结构，达到了 1.85 Å 的分辨率，这是一种产物类似物。该结构表明它属于 cupin 超家族。在存在 NADPH 的情况下，解决了 Cj1428 的三维晶体结构，分辨率为 1.50 Å。它的折叠使其归入短链脱氢酶/还原酶超家族。通常，该家族中的成员显示出 YXXXK 的特征签名序列，其中保守的酪氨酸在催化中起关键作用。在 Cj1428 中，该残基是苯丙氨酸。
