Plant peptidoglycan precursor biosynthesis: Conservation between moss chloroplasts and Gram negative bacteria
Plant peptidoglycan precursor biosynthesis: Conservation between moss chloroplasts and Gram negative bacteria
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植物肽聚糖前体生物合成：苔藓叶绿体和革兰氏阴性细菌之间的保守性
DOI：
10.1101/2022.01.05.475093
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发表时间：
2022
期刊：
影响因子：
--
通讯作者：
Dowson A
中科院分区：
文献类型：
--
作者：
Dowson A
越来越多的证据表明，在许多光合作用真核生物的叶绿体周围合成了与细菌细胞壁一致的肽聚糖，从蓝藻到包括蕨类植物在内的早期分叉的陆地植物，但其生物合成途径尚未被证实。在这里，我们使用了质谱学和酶学的双重方法来表征苔藓(Physcomitrium(Physcomitrella)patens)叶绿体中肽多糖的合成。为了促进肽聚糖途径中间体的积累，用抗生素磷霉素、D-环丝氨酸和羧苯西林进行了培养，这些抗生素抑制了细菌中关键的肽聚糖途径蛋白。经三氯乙酸提取的苔藓代谢物的质谱分析显示，尿苷二磷酸N-乙酰氨基葡萄糖(UDP-GlcNAc)通过尿苷二磷酸-N-乙酰壁氨酸(UDP-MurNAc)-D，L-二氨基戊二酸酯(DAP)-五肽的五个预测中间体的水平升高。大多数革兰氏阴性细菌，包括蓝藻，都将异二氨基苯二酸(D，L-DAP)结合到肽聚糖的茎肽的第三个残基中，而不是大多数革兰氏阳性细菌典型的L赖氨酸。目的：确定D，L-DAP掺入TEP的特异性。在此基础上，我们对两种菌株的重组蛋白UDP-N-acetylmuramoyl-L-alanyl-D-glutamate–2，6-diaminopimelate连接酶(Mure)进行了分析。专利和蓝藻鱼腥藻(Nostoc sp.菌株PCC7120)。这两种连接酶几乎完全优先结合D，L-DAP，而不是L-赖氨酸，这与质谱学数据一致，其催化效率与先前报道的革兰氏阴性细菌Mure连接酶相似。我们讨论了这些数据如何符合DL-DAP-掺入细菌Mure连接酶共同的活性位点残基的保守性，以及植物肽聚糖途径中水平基因转移事件的可能性。
Accumulating evidence suggests that peptidoglycan, consistent with a bacterial cell wall, is synthesized around the chloroplasts of many photosynthetic eukaryotes, from glaucophyte algae to early-diverging land plants including pteridophyte ferns, but the biosynthetic pathway has not been demonstrated. Here, we employed mass spectrometry and enzymology in a two-fold approach to characterize the synthesis of peptidoglycan in chloroplasts of the mossPhyscomitrium(Physcomitrella)patens. To drive the accumulation of peptidoglycan pathway intermediates,P. patenswas cultured with the antibiotics fosfomycin, D-cycloserine, and carbenicillin, which inhibit key peptidoglycan pathway proteins in bacteria. Mass spectrometry of the trichloroacetic acid-extracted moss metabolome revealed elevated levels of five of the predicted intermediates from uridine diphosphateN-acetylglucosamine (UDP-GlcNAc) through the uridine diphosphateN-acetylmuramic acid (UDP-MurNAc)-D,L-diaminopimelate (DAP)-pentapeptide. Most Gram-negative bacteria, including cyanobacteria, incorporatemeso-diaminopimelic acid (D,L-DAP) into the third residue of the stem peptide of peptidoglycan, as opposed to L-lysine, typical of most Gram-positive bacteria. To establish the specificity of D,L-DAP incorporation into theP. patensprecursors, we analyzed the recombinant protein UDP-N-acetylmuramoyl-L-alanyl-D-glutamate–2,6-diaminopimelate ligase (MurE) from bothP. patensand the cyanobacteriumAnabaena sp.(Nostoc sp.strain PCC 7120). Both ligases incorporated D,L-DAP in almost complete preference to L-Lys, consistent with the mass spectrophotometric data, with catalytic efficiencies similar to previously documented Gram-negative bacterial MurE ligases. We discuss how these data accord with the conservation of active site residues common to DL-DAP-incorporating bacterial MurE ligases and of the probability of a horizontal gene transfer event within the plant peptidoglycan pathway.