Suzuki Nao, Nakano Yoshio, Yoshida Yasuo, Nezu Takashi, Terada Yoshihiro, Yamashita Yoshihisa, Koga Toshihiko
Department of Preventive Dentistry, Kyushu University Faculty of Dental Science, Fukuoka, Japan.
Eur J Biochem. 2002 Dec;269(23):5963-71. doi: 10.1046/j.1432-1033.2002.03331.x.
The serotype a-specific polysaccharide antigen of Actinobacillus actinomycetemcomitans is an unusual sugar, 6-deoxy-d-talose. Guanosine diphosphate (GDP)-6-deoxy-d-talose is the activated sugar nucleotide form of 6-deoxy-d-talose, which has been identified as a constituent of only a few microbial polysaccharides. In this paper, we identify two genes encoding GDP-6-deoxy-d-talose synthetic enzymes, GDP-alpha-d-mannose 4,6-dehydratase and GDP-4-keto-6-deoxy-d-mannose reductase, in the gene cluster required for the biosynthesis of serotype a-specific polysaccharide antigen from A. actinomycetemcomitans SUNYaB 75. Both gene products were produced and purified from Escherichia coli transformed with plasmids containing these genes. Their enzymatic reactants were analysed by reversed-phase HPLC (RP-HPLC). The sugar nucleotide produced from GDP-alpha-d-mannose by these enzymes was purified by RP-HPLC and identified by electrospray ionization-MS, 1H nuclear magnetic resonance, and GC/MS. The results indicated that GDP-6-deoxy-d-talose is produced from GDP-alpha-d-mannose. This paper is the first report on the GDP-6-deoxy-d-talose biosynthetic pathway and the role of GDP-4-keto-6-deoxy-d-mannose reductase in the synthesis of GDP-6-deoxy-d-talose.
伴放线放线杆菌a血清型特异性多糖抗原是一种特殊的糖类，即6-脱氧-D-塔罗糖。鸟苷二磷酸（GDP）-6-脱氧-D-塔罗糖是6-脱氧-D-塔罗糖的活性糖核苷酸形式，仅在少数微生物多糖中被鉴定为一种成分。在本文中，我们在伴放线放线杆菌SUNYaB 75 a血清型特异性多糖抗原生物合成所需的基因簇中，鉴定出两个编码GDP-6-脱氧-D-塔罗糖合成酶的基因，即GDP-α-D-甘露糖4,6-脱水酶和GDP-4-酮-6-脱氧-D-甘露糖还原酶。这两种基因产物都是从用含有这些基因的质粒转化的大肠杆菌中产生并纯化的。通过反相高效液相色谱（RP-HPLC）分析它们的酶促反应物。通过RP-HPLC纯化这些酶从GDP-α-D-甘露糖产生的糖核苷酸，并通过电喷雾电离质谱、1H核磁共振和气相色谱/质谱进行鉴定。结果表明，GDP-6-脱氧-D-塔罗糖是由GDP-α-D-甘露糖产生的。本文首次报道了GDP-6-脱氧-D-塔罗糖的生物合成途径以及GDP-4-酮-6-脱氧-D-甘露糖还原酶在GDP-6-脱氧-D-塔罗糖合成中的作用。
