Characterization of serine hydroxymethyltransferase GlyA as a potential source of D-alanine in Chlamydia pneumoniae
Characterization of serine hydroxymethyltransferase GlyA as a potential source of D-alanine in Chlamydia pneumoniae
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DOI：
10.3389/fcimb.2014.00019
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发表时间：
2014-02-01
影响因子：
5.7
通讯作者：
Henrichfreise, Beate
中科院分区：
文献类型：
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作者：
De Benedetti, Stefania;Buehl, Henrike;Henrichfreise, Beate
对于细胞内衣原体科，不需要承受渗透挑战，到目前为止还没有在这些病原体中检测到功能细胞壁。然而，青霉素抑制衣原体科的细胞分裂，导致扩大的异常体，这种现象被称为衣原体异常。d -丙氨酸是细菌细胞壁生物合成中独特而必需的成分。在像大肠杆菌这样的自由生活细菌中，青霉素结合蛋白，如单功能转肽酶PBP2和PBP3（衣原体科中青霉素的假定靶点），通过五肽侧链的中二氨基戊二酸和D-Ala-D-Ala部分，将相邻的肽聚糖链交联。由于缺乏编码丙氨酸消旋酶Alr和DadX同源基因的基因，衣原科中D-Ala的来源以及PBP2和PBP3活性底物的存在多年来一直困扰着研究人员。有趣的是，衣原体科基因组编码GlyA，这是一种丝氨酸羟甲基转移酶，在大肠杆菌中显示出D-和l -丙氨酸的缓慢外消旋反应。我们发现来自肺炎衣原体的GlyA可以作为D-Ala的来源。GlyA部分逆转了大肠杆菌消旋酶双突变体的D-Ala营养不良表型。此外，纯化的衣原体GlyA在体外对L-Ala具有消旋酶活性，并被d -环丝氨酸抑制，这表明GlyA除了D-Ala连接酶MurC/Ddl外，是该竞争性抑制剂在衣原体科中的另一个靶点。衣原体科中D-Ala生物合成的证据有助于阐明细胞壁前体脂质II的结构以及衣原体青霉素结合蛋白在非分裂异常衣原体的发育和青霉素存在下的持久性中的作用。
For intracellular Chlamydiaceae, there is no need to withstand osmotic challenges, and a functional cell wall has not been detected in these pathogens so far. Nevertheless, penicillin inhibits cell division in Chlamydiaceae resulting in enlarged aberrant bodies, a phenomenon known as chlamydial anomaly. D-alanine is a unique and essential component in the biosynthesis of bacterial cell walls. In free-living bacteria like Escherichia coli, penicillin-binding proteins such as monofunctional transpeptidases PBP2 and PBP3, the putative targets of penicillin in Chlamydiaceae, cross-link adjacent peptidoglycan strands via meso-diaminopimelic acid and D-Ala-D-Ala moieties of pentapeptide side chains. In the absence of genes coding for alanine racemase Alr and DadX homologs, the source of D-Ala and thus the presence of substrates for PBP2 and PBP3 activity in Chlamydiaceae has puzzled researchers for years. Interestingly, Chlamydiaceae genomes encode GlyA, a serine hydroxymethyltransferase that has been shown to exhibit slow racemization of D- and L-alanine as a side reaction in E. coli. We show that GlyA from Chlamydia pneumoniae can serve as a source of D-Ala. GlyA partially reversed the D-Ala auxotrophic phenotype of an E. coli racemase double mutant. Moreover, purified chlamydial GlyA had racemase activity on L-Ala in vitro and was inhibited by D-cycloserine, identifying GlyA, besides D-Ala ligase MurC/Ddl, as an additional target of this competitive inhibitor in Chlamydiaceae. Proof of D-Ala biosynthesis in Chlamydiaceae helps to clarify the structure of cell wall precursor lipid II and the role of chlamydial penicillin-binding proteins in the development of non-dividing aberrant chlamydial bodies and persistence in the presence of penicillin.