Selective penicillin-binding protein imaging probes reveal substructure in bacterial cell division.
Selective penicillin-binding protein imaging probes reveal substructure in bacterial cell division.
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DOI：
10.1021/cb300329r
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发表时间：
2012-10-19
影响因子：
4
通讯作者：
Carlson, Erin E.
中科院分区：
文献类型：
--
作者：
Kocaoglu, Ozden;Calvo, Rebecca A.;Sham, Lok-To;Cozy, Loralyn M.;Lanning, Bryan R.;Francis, Samson;Winkler, Malcolm E.;Kearns, Daniel B.;Carlson, Erin E.
肽聚糖细胞壁是抗生素治疗的常见靶标，但其结构和组装仅部分了解。肽聚糖的合成需要一套青霉素结合蛋白（PBP），其各自的作用是难以确定的，因为每种酶往往是生长缓慢，可能是由于功能冗余。为了应对这一挑战，我们试图生成能够对PBPs子集进行选择性检查的工具。我们设计并合成了含β-内酰胺抗生素头孢菌素C的荧光衍生物和生物素衍生物。这些探针促进了枯草芽孢杆菌PY 79和D39肺炎链球菌的未包封衍生物中活性PBPs的特异性体内标记。显微镜和基于凝胶的分析表明，头孢菌素C为基础的探针比BOCILLIN-FL，一种市售的青霉素V类似物，标记所有的PBPs更具选择性。通过饱和头孢菌素C敏感的PBP，然后用BOCILLIN-FL标记剩余的PBP群体来进行活细胞的双重标记，证明两组PBP不共定位。这表明，即使是位于特定位点的PBP（例如，隔膜）并不都是混合的，而是PBP亚群离散本地化。因此，Ceph C探针代表了探索PBP子集的新工具，并有可能促进对这类关键蛋白质作用的更深入了解。
The peptidoglycan cell wall is a common target for antibiotic therapy but its structure and assembly are only partially understood. Peptidoglycan synthesis requires a suite of penicillin-binding proteins (PBPs), the individual roles of which are difficult to determine because each enzyme is often dispensable for growth perhaps due to functional redundancy. To address this challenge, we sought to generate tools that would enable selective examination of a subset of PBPs. We designed and synthesized fluorescent and biotin derivatives of the β-lactam-containing antibiotic cephalosporin C. These probes facilitated specific in vivo labeling of active PBPs in both Bacillus subtilis PY79 and an unencapsulated derivative of D39 Streptococcus pneumoniae. Microscopy and gel-based analysis indicated that the cephalosporin C-based probes are more selective than BOCILLIN-FL, a commercially available penicillin V analog, which labels all PBPs. Dual labeling of live cells performed by saturation of cephalosporin C-susceptible PBPs followed by tagging of the remaining PBP population with BOCILLIN-FL demonstrated that the two sets of PBPs are not co-localized. This suggests that even PBPs that are located at a particular site (e.g., septum) are not all intermixed, but rather that PBP subpopulations are discretely localized. Accordingly, the Ceph C probes represent new tools to explore a subset of PBPs and have the potential to facilitate a deeper understand of the roles of this critical class of proteins.
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影响因子：
64.8
作者：
SPRATT, BG;PARDEE, AB
通讯作者：
PARDEE, AB
影响因子：
3.2
作者：
Barendt, Skye M.;Sham, Lok-To;Winkler, Malcolm E.
通讯作者：
Winkler, Malcolm E.
影响因子：
64.5
作者：
Paradis-Bleau C;Markovski M;Uehara T;Lupoli TJ;Walker S;Kahne DE;Bernhardt TG
通讯作者：
Bernhardt TG
影响因子：
3.2
作者：
Land, Adrian D.;Winkler, Malcolm E.
通讯作者：
Winkler, Malcolm E.
影响因子：
3.2
作者：
McPherson, DC;Driks, A;Popham, DL
通讯作者：
Popham, DL