Amino Acid Substitutions of MagA in Klebsiella pneumoniae Affect the Biosynthesis of the Capsular Polysaccharide
Amino Acid Substitutions of MagA in Klebsiella pneumoniae Affect the Biosynthesis of the Capsular Polysaccharide
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DOI：
10.1371/journal.pone.0046783
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发表时间：
2012-10-31
期刊：
影响因子：
3.7
通讯作者：
Wang, Jin-Town
中科院分区：
文献类型：
--
作者：
Lin, Tzu-Lung;Yang, Feng-Ling;Wang, Jin-Town
肺炎克雷伯氏菌的粘膜粘度相关基因A（magA）参与K1荚膜多糖（CPS）的生物合成。基于序列同源性和基因比对，已预测magA基因编码Wzy型CPS聚合酶。与Wzy_C和RfaL蛋白家族（其催化CPS或脂多糖（LPS）生物合成）的序列比对和拓扑分析表明，包括G308、G310、G334、G337、R290、P305、H323和N324在内的8个高度保守的残基位于假设的环区域中。因此，我们使用定点诱变来研究这些残基在CPS生产中的作用，并观察化脓性肝脓肿菌株NTUH-K2044中随后的表型，如粘膜粘度、血清和吞噬抗性以及毒力（如在小鼠中评估的）。在R290或H323处的丙氨酸取代废除了所有这些性质。G308 A突变体的这些功能严重受损。G334 A突变体保持粘液样，CPS产量减少，但其体内毒力显著降低。携带magA G310 A、G337 A、P305 A或N324 A突变的菌株未观察到表型变化。因此，R290、G308、H323和G334是克雷伯氏菌MagA（Wzy）蛋白的功能重要残基。pneumoniae NTUH-K2044，荚膜型K1。这些氨基酸也可能是重要的功能，Wzy在其他荚膜类型的K。pneumoniae和其他携带Wzy_C家族蛋白的物种。
Mucoviscosity-associated gene A (magA) of Klebsiella pneumoniae contributes to K1 capsular polysaccharide (CPS) biosynthesis. Based on sequence homology and gene alignment, the magA gene has been predicted to encode a Wzy-type CPS polymerase. Sequence alignment with the Wzy_C and RfaL protein families (which catalyze CPS or lipopolysaccharide (LPS) biosynthesis) and topological analysis has suggested that eight highly conserved residues, including G308, G310, G334, G337, R290, P305, H323, and N324, were located in a hypothetical loop region. Therefore, we used site-directed mutagenesis to study the role of these residues in CPS production, and to observe the consequent phenotypes such as mucoviscosity, serum and phagocytosis resistance, and virulence (as assessed in mice) in pyogenic liver abscess strain NTUH-K2044. Alanine substitutions at R290 or H323 abolished all of these properties. The G308A mutant was severely impaired for these functions. The G334A mutant remained mucoid with decreased CPS production, but its virulence was significantly reduced in vivo. No phenotypic change was observed for strains harboring magA G310A, G337A, P305A, or N324A mutations. Therefore, R290, G308, H323, and G334 are functionally important residues of the MagA (Wzy) protein of K. pneumoniae NTUH-K2044, capsular type K1. These amino acids are also likely to be important for the function of Wzy in other capsular types in K. pneumoniae and other species bearing Wzy_C family proteins.