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苜蓿中华根瘤菌中磷脂N-甲基转移酶基因的失活:正常生长需要磷脂酰胆碱。

Inactivation of the gene for phospholipid N-methyltransferase in Sinorhizobium meliloti: phosphatidylcholine is required for normal growth.

作者信息

de Rudder K E, López-Lara I M, Geiger O

机构信息

Institute of Biotechnology, Technical University Berlin, Germany.

出版信息

Mol Microbiol. 2000 Aug;37(4):763-72. doi: 10.1046/j.1365-2958.2000.02032.x.

DOI:10.1046/j.1365-2958.2000.02032.x
PMID:10972799
Abstract

In phosphatidylcholine (PC)-containing prokaryotes, only the methylation pathway of PC biosynthesis was thought to occur. However, a second choline-dependent pathway for PC formation, the PC synthase (Pcs) pathway, exists in Sinorhizobium (Rhizobium) meliloti in which choline is condensed with CDP-diacylglyceride. Here, we characterize the methylation pathway of PC biosynthesis in S. meliloti. A mutant deficient in phospholipid N-methyltransferase (Pmt) was complemented with a S. meliloti gene bank and the complementing DNA was sequenced. A gene coding for a S-adenosylmethionine-dependent N-methyltransferase was identified as the sinorhizobial Pmt, which showed little similarity to the corresponding enzyme from Rhodobacter sphaeroides. Upon expression of the sinorhizobial Pmt, besides phosphatidylcholine, the methylated intermediates of the methylation pathway, monomethylphosphatidylethanolamine and dimethylphosphatidylethanolamine, are also formed. When Pmt-deficient mutants of S. meliloti are grown on minimal medium, they cannot form PC, and they grow significantly more slowly than the wild type. Growth of the Pmt-deficient mutant in the presence of choline allows for PC formation via the Pcs pathway and restores wild-type-like growth. Double knock-out mutants, deficient in Pmt and in Pcs, are unable to form PC and show reduced growth even in the presence of choline. These results suggest that PC is required for normal growth of S. meliloti.

摘要

在含有磷脂酰胆碱(PC)的原核生物中,人们曾认为仅存在PC生物合成的甲基化途径。然而,在苜蓿中华根瘤菌(Sinorhizobium (Rhizobium) meliloti)中存在第二条依赖胆碱的PC形成途径,即PC合酶(Pcs)途径,其中胆碱与CDP - 二酰甘油缩合。在此,我们对苜蓿中华根瘤菌中PC生物合成的甲基化途径进行了表征。一个磷脂N - 甲基转移酶(Pmt)缺陷型突变体用苜蓿中华根瘤菌基因文库进行互补,并对互补DNA进行测序。一个编码依赖S - 腺苷甲硫氨酸的N - 甲基转移酶的基因被鉴定为中华根瘤菌Pmt,它与球形红杆菌(Rhodobacter sphaeroides)的相应酶几乎没有相似性。在表达中华根瘤菌Pmt后,除了磷脂酰胆碱外,甲基化途径的甲基化中间体单甲基磷脂酰乙醇胺和二甲基磷脂酰乙醇胺也会形成。当苜蓿中华根瘤菌的Pmt缺陷型突变体在基本培养基上生长时,它们无法形成PC,并且生长速度明显比野生型慢得多。在胆碱存在的情况下,Pmt缺陷型突变体的生长允许通过Pcs途径形成PC,并恢复类似野生型的生长。Pmt和Pcs双敲除突变体即使在胆碱存在的情况下也无法形成PC,并且生长受到抑制。这些结果表明PC是苜蓿中华根瘤菌正常生长所必需的。

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