Properties of a novel PBP2A protein homolog from Staphylococcus aureus strain LGA251 and its contribution to the β-lactam-resistant phenotype.
Properties of a novel PBP2A protein homolog from Staphylococcus aureus strain LGA251 and its contribution to the β-lactam-resistant phenotype.
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DOI：
10.1074/jbc.m112.395962
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发表时间：
2012-10-26
期刊：
影响因子：
--
通讯作者：
Tomasz A
中科院分区：
文献类型：
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作者：
Kim C;Milheiriço C;Gardete S;Holmes MA;Holden MT;de Lencastre H;Tomasz A
背景：最近分离出的MRSA LGA251具有低耐药性，并携带新的mecA同源物。结果：新 mecA 的蛋白质产物 PBP2ALGA 显示出对青霉素的“偏好”，并且在 37 °C 下不稳定。 mecALGA251 引入易感金黄色葡萄球菌后可表达高水平耐药性。意义：本研究深入了解 PBP2A 样蛋白的结构和功能之间的关系。耐甲氧西林金黄色葡萄球菌 (MRSA) 菌株在耐药水平、遗传背景以及携带耐药基因 mecA 的染色体盒 (SCCmec) 结构方面表现出菌株间差异。相比之下，在迄今为止检测的 MRSA 分离株中，发现 mecA 决定簇序列的菌株间变异要有限得多。第一个例外是最近鉴定出的 MRSA 菌株 LGA251，它携带该基因的新同源物以及同样具有新颖、高度差异结构的调控元件 mecI/mecR。在大肠杆菌中克隆和纯化后，新 mecA 同源物的蛋白质产物 PBP2ALGA 在 SDS-PAGE 中显示出异常的迁移率、结构不稳定且在 37 °C 下失去活性，并且与头孢西丁相比对苯唑西林具有更高的相对亲和力。将不含调控元件的 mecA 同源物克隆到质粒中，并引入到 β-内酰胺敏感的金黄色葡萄球菌菌株 COL-S 的背景中。在此背景下，mecA 同源物表现出对头孢西丁的高水平耐药性（MIC = 400 μg/ml）和对苯唑西林的较低耐药性（最低抑制浓度 = 200 μg/ml）。与 PBP2A 类似，蛋白质同源物 PBP2ALGA 能够替代金黄色葡萄球菌 PBP2 生长的基本功能。与 PBP2A 相比，PBP2ALGA 不依赖于天然 PBP2 的转糖基酶活性来表达对苯唑西林的高水平抗性，这表明 PBP2A 同源物可能优先与单功能转糖基酶合作作为转糖基酶活性的替代来源。
Background: The recently isolated MRSA LGA251 has low resistance and carries a new mecA homolog. Results: PBP2ALGA, the protein product of the new mecA, showed a “preference” for penicillins and instability at 37 °C. mecALGA251 introduced into susceptible S. aureus allowed expression of high-level resistance. Significance: This study provides insights into the relationship between structure and function of PBP2A-like proteins. Methicillin-resistant Staphylococcus aureus (MRSA) strains show strain-to-strain variation in resistance level, in genetic background, and also in the structure of the chromosomal cassette (SCCmec) that carries the resistance gene mecA. In contrast, strain-to-strain variation in the sequence of the mecA determinant was found to be much more limited among MRSA isolates examined so far. The first exception to this came with the recent identification of MRSA strain LGA251, which carries a new homolog of this gene together with regulatory elements mecI/mecR that also have novel, highly divergent structures. After cloning and purification in Escherichia coli, PBP2ALGA, the protein product of the new mecA homolog, showed aberrant mobility in SDS-PAGE, structural instability and loss of activity at 37 °C, and a higher relative affinity for oxacillin as compared with cefoxitin. The mecA homolog free of its regulatory elements was cloned into a plasmid and introduced into the background of the β-lactam-susceptible S. aureus strain COL-S. In this background, the mecA homolog expressed a high-level resistance to cefoxitin (MIC = 400 μg/ml) and a somewhat lower resistance to oxacillin (minimal inhibitory concentration = 200 μg/ml). Similar to PBP2A, the protein homolog PBP2ALGA was able to replace the essential function of the S. aureus PBP2 for growth. In contrast to PBP2A, PBP2ALGA did not depend on the transglycosylase activity of the native PBP2 for expression of high level resistance to oxacillin, suggesting that the PBP2A homolog may preferentially cooperate with a monofunctional transglycosylase as the alternative source of transglycosylase activity.