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首页> 外文期刊>Infection and immunity >Analysis of the Specificity of Panton-Valentine Leucocidin and Gamma-Hemolysin F Component Binding
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Analysis of the Specificity of Panton-Valentine Leucocidin and Gamma-Hemolysin F Component Binding

机译：Panton-Valentine Leucocidin和γ-溶血素F成分结合的特异性分析

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摘要

In this study, the binding of F components of the staphylococcal bicomponent leukotoxins Panton-Valentine leucocidin (LukF-PV) and gamma-hemolysin (HlgB) on polymorphonuclear neutrophils (PMNs), monocytes, and lymphocytes was determined using labeled mutants and flow cytometry. Leukotoxin activity was evaluated by measuring Ca2+ entry or pore formation using spectrofluorometry or flow cytometry. Although HlgB had no affinity for cells in the absence of an S component, LukF-PV had high affinity for PMNs (dissociation constant [Kd], 6.2 ± 1.9 nM; n = 8), monocytes (Kd, 2.8 ± 0.8 nM; n = 7), and lymphocytes (Kd, 1.2 ± 0.2 nM; n = 7). Specific binding of HlgB was observed only after addition of LukS-PV on PMNs (Kd, 1.1 ± 0.2 nM; n = 4) and monocytes (Kd, 0.84 ± 0.31 nM; n = 4) or after addition of HlgC on PMNs, monocytes, and lymphocytes. Addition of LukS-PV or HlgC induced a second specific binding of LukF-PV on PMNs. HlgB and LukD competed only with LukF-PV molecules bound after addition of LukS-PV. LukF-PV and LukD competed with HlgB in the presence of LukS-PV on PMNs and monocytes. Use of antibodies and comparisons between binding and activity time courses showed that the LukF-PV molecules that bound to target cells before addition of LukS-PV were the only LukF-PV molecules responsible for Ca2+ entry and pore formation. In contrast, the active HlgB molecules were the HlgB molecules bound after addition of LukS-PV. In conclusion, LukF-PV must be linked to LukS-PV and to a binding site of the membrane to have toxin activity.
机译：在这项研究中，使用标记的突变体和流式细胞仪确定了葡萄球菌双组分白细胞毒素F组分Panton-Valentine leucocidin（LukF-PV）和γ-溶血素（HlgB）在多形核中性粒细胞（PMN），单核细胞和淋巴细胞上的结合。通过使用荧光分光光度法或流式细胞术测量Ca 2 + 的进入或孔形成来评估白细胞毒素活性。尽管在没有S组分的情况下HlgB对细胞没有亲和力，但LukF-PV对PMN具有高亲和力（解离常数[ K d ]，6.2±1.9 nM; n = 8），单核细胞（ K d ，2.8±0.8 nM; n = 7）和淋巴细胞（ K d ，1.2±0.2 nM; n = 7）。仅在将LukS-PV加入PMN后观察到HlgB的特异性结合（ K d ，1.1±0.2 nM; n < / em> = 4）和单核细胞（ K d ，0.84±0.31 nM; n = 4）或在PMN，单核细胞和淋巴细胞上添加HlgC之后。 LukS-PV或HlgC的添加诱导了LukF-PV对PMN的第二种特异性结合。 HlgB和LukD仅与加入LukS-PV后结合的LukF-PV分子竞争。在存在LukS-PV的情况下，LukF-PV和LukD与HlgB在PMN和单核细胞上竞争。抗体的使用以及结合和活动时间过程的比较表明，在添加LukS-PV之前与靶细胞结合的LukF-PV分子是唯一负责Ca 2 + 进入和转移的LukF-PV分子。孔形成。相反，活性HlgB分子是添加LukS-PV后结合的HlgB分子。总之，LukF-PV必须与LukS-PV和膜的结合位点连接才能具有毒素活性。

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