Ranjit Dev K, Jorgenson Matthew A, Young Kevin D
Department of Microbiology and Immunology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA.
Department of Microbiology and Immunology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA
J Bacteriol. 2017 Mar 14;199(7). doi: 10.1128/JB.00612-16. Print 2017 Apr 1.
Peptidoglycan is a vital component of nearly all cell wall-bearing bacteria and is a valuable target for antibacterial therapy. However, despite decades of work, there remain important gaps in understanding how this macromolecule is synthesized and molded into a three-dimensional structure that imparts specific morphologies to individual cells. Here, we investigated the particularly enigmatic area of how peptidoglycan is synthesized and shaped during the first stages of creating cell shape , that is, in the absence of a preexisting template. We found that when lysozyme-induced (LI) spheroplasts of were allowed to resynthesize peptidoglycan, the cells divided first and then elongated to recreate a normal rod-shaped morphology. Penicillin binding protein 1B (PBP1B) was critical for the first stage of this recovery process. PBP1B synthesized peptidoglycan , and this synthesis required that PBP1B interact with the outer membrane lipoprotein LpoB. Surprisingly, when LpoB was localized improperly to the inner membrane, recovering spheroplasts synthesized peptidoglycan and divided but then propagated as amorphous spheroidal cells, suggesting that the regeneration of a normal rod shape depends on a particular spatial interaction. Similarly, spheroplasts carrying a PBP1B variant lacking transpeptidase activity or those in which PBP1A was overproduced could synthesize new peptidoglycan and divide but then grew as oddly shaped spheroids. We conclude that cell wall synthesis requires the glycosyltransferase activity of PBP1B but that PBP1B transpeptidase activity is needed to assemble cell walls with wild-type morphology. Bacterial cell wall peptidoglycan is synthesized and modified by penicillin binding proteins (PBPs), which are targeted by about half of all currently prescribed antibiotics, including penicillin and its derivatives. Because antibiotic resistance is rising, it has become increasingly urgent that we fill the gaps in our knowledge about how PBPs create and assemble this protective wall. We report here that PBP1B plays an essential role in synthesizing peptidoglycan in the absence of a preexisting template: its glycosyltransferase activity is responsible for synthesis, while its transpeptidase activity is required to construct cell walls of a specific shape. These results highlight the importance of this enzyme and distinguish its biological roles from those of other PBPs and peptidoglycan synthases.
肽聚糖是几乎所有具有细胞壁的细菌的重要组成部分，也是抗菌治疗的一个重要靶点。然而，尽管经过了数十年的研究，但在理解这种大分子如何合成并塑造成赋予单个细胞特定形态的三维结构方面，仍存在重大差距。在这里，我们研究了肽聚糖在细胞形状形成的最初阶段，即在没有预先存在模板的情况下，是如何合成和塑造的这一特别神秘的领域。我们发现，当溶菌酶诱导的（LI）球形芽孢杆菌原生质体被允许重新合成肽聚糖时，细胞首先分裂，然后伸长以重新形成正常的杆状形态。青霉素结合蛋白1B（PBP1B）对这一恢复过程的第一阶段至关重要。PBP1B合成肽聚糖，而这种合成要求PBP1B与外膜脂蛋白LpoB相互作用。令人惊讶的是，当LpoB在内膜上定位不当，恢复的原生质体合成肽聚糖并分裂，但随后以无定形球状细胞的形式繁殖，这表明正常杆状形态的再生取决于特定的空间相互作用。同样，携带缺乏转肽酶活性的PBP1B变体的原生质体或过量产生PBP1A的原生质体可以合成新的肽聚糖并分裂，但随后长成形状怪异的球体。我们得出结论，球形芽孢杆菌细胞壁合成需要PBP1B的糖基转移酶活性，但需要PBP1B的转肽酶活性来组装具有野生型形态的细胞壁。细菌细胞壁肽聚糖由青霉素结合蛋白（PBPs）合成和修饰，目前约一半的处方抗生素，包括青霉素及其衍生物，都以PBPs为靶点。由于抗生素耐药性不断上升，填补我们对PBPs如何构建和组装这一保护壁的知识空白变得越来越紧迫。我们在此报告，PBP1B在没有预先存在模板的情况下，在合成肽聚糖中起重要作用：其糖基转移酶活性负责肽聚糖的合成，而其转肽酶活性是构建特定形状细胞壁所必需的。这些结果突出了这种酶的重要性，并将其生物学作用与其他PBPs和肽聚糖合酶的作用区分开来。
