Lee Jeongmi, Sperandio Vanessa, Frantz Doug E, Longgood Jamie, Camilli Andrew, Phillips Margaret A, Michael Anthony J
Departments of Pharmacology, Microbiology, and Biochemistry, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas 75390, USA.
J Biol Chem. 2009 Apr 10;284(15):9899-907. doi: 10.1074/jbc.M900110200. Epub 2009 Feb 5.
Polyamines are small organic cations found in all cells, and the biosynthetic pathway is well described in eukaryotes and Escherichia coli. The characterized pathway uses decarboxylated S-adenosylmethionine as the aminopropyl group donor to form spermidine from putrescine by the key enzymes S-adenosylmethionine decarboxylase and spermidine synthase. We report here the in vivo characterization of an alternative polyamine biosynthetic pathway from Vibrio cholerae, the causative agent of human cholera. The pathway uses aspartate beta-semialdehyde as the aminopropyl group donor and consists of a fused protein containing l-2,4-diaminobutyrate aminotransferase and l-2,4-diaminobutyrate decarboxylase, a carboxynorspermidine dehydrogenase (CANSDH), and a carboxynorspermidine decarboxylase (CANSDC). We show that in V. cholerae, this pathway is required for synthesis of both sym-norspermidine and spermidine. Heterologous expression of the V. cholerae pathway in E. coli results in accumulation of the nonnative polyamines diaminopropane and sym-norspermidine. Genetic deletion of the V. cholerae CANSDC led to accumulation of carboxynorspermidine, whereas deletion of either CANSDC or the putative CANSDH led to loss of sym-norspermidine and spermidine. These results allowed unambiguous identification of the gene encoding CANSDH. Furthermore, deletion of either CANSDH or CANSDC led to a 50-60% reduction in growth rate of planktonic cells and severely reduced biofilm formation, which could be rescued by exogenously supplied sym-norspermidine but not spermidine. The pathway was not required for infectivity in a mouse model of V. cholerae infection. Notably, the alternative polyamine biosynthetic pathway is widespread in bacteria and is likely to play a previously unrecognized role in the biology of these organisms.
多胺是存在于所有细胞中的小分子有机阳离子，其生物合成途径在真核生物和大肠杆菌中已有详尽描述。该特征性途径利用脱羧的S - 腺苷甲硫氨酸作为氨丙基供体，通过关键酶S - 腺苷甲硫氨酸脱羧酶和亚精胺合酶，由腐胺形成亚精胺。我们在此报告了霍乱弧菌（人类霍乱的病原体）另一条多胺生物合成途径的体内特征。该途径使用天冬氨酸β - 半醛作为氨丙基供体，由包含L - 2,4 - 二氨基丁酸转氨酶和L - 2,4 - 二氨基丁酸脱羧酶的融合蛋白、羧基去甲亚精胺脱氢酶（CANSDH）和羧基去甲亚精胺脱羧酶（CANSDC）组成。我们表明，在霍乱弧菌中，该途径对于合成对称去甲亚精胺和亚精胺都是必需的。霍乱弧菌途径在大肠杆菌中的异源表达导致非天然多胺二氨基丙烷和对称去甲亚精胺的积累。霍乱弧菌CANSDC的基因缺失导致羧基去甲亚精胺的积累，而CANSDC或假定的CANSDH的缺失导致对称去甲亚精胺和亚精胺的缺失。这些结果使得能够明确鉴定编码CANSDH的基因。此外，CANSDH或CANSDC的缺失导致浮游细胞生长速率降低50 - 60%，并严重减少生物膜形成，外源性提供的对称去甲亚精胺而非亚精胺可挽救这种情况。在霍乱弧菌感染的小鼠模型中，该途径对于感染性不是必需的。值得注意的是，这条替代的多胺生物合成途径在细菌中广泛存在，并且可能在这些生物体的生物学中发挥以前未被认识到的作用。
