Repeat regions R1 and R2 in the P97 paralogue Mhp271 of Mycoplasma hyopneumoniae bind heparin, fibronectin and porcine cilia
Repeat regions R1 and R2 in the P97 paralogue Mhp271 of Mycoplasma hyopneumoniae bind heparin, fibronectin and porcine cilia
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DOI：
10.1111/j.1365-2958.2010.07345.x
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发表时间：
2010-10-01
影响因子：
3.6
通讯作者：
Djordjevic, Steven P.
中科院分区：
文献类型：
--
作者：
Deutscher, Ania T.;Jenkins, Cheryl;Djordjevic, Steven P.
猪肺炎支原体是引起猪地方性肺炎的病原体，它附着在有纤毛的呼吸道上皮细胞上，导致纤毛淤滞和上皮细胞死亡。纤毛粘附素P97(Mhp183)包含两个重复区域，分别命名为R1和R2，它们在粘附性中发挥关键作用。R1中的8个五肽重复足以结合猪纤毛；然而，R1和R2都需要结合肝素。Mhp271是P97的同源蛋白，是唯一一种同时含有R1和R2重复序列的猪肺炎支原体蛋白。这些重复序列被安排为一组三个五肽重复序列(标记为R1a(271))、两个十肽重复序列(标记为R2(271))和第二组六个五肽重复序列(标记为R1b(271))。为了确定它们的功能，构建了含有R1a(271)(F1(271))和R2(271)-R1b(271)(F2(271))的重组蛋白，并用于体外结合试验。F2(271)，但不包括F1(271)、结合肝素(K-D=8.1+/-0.4 nM)、纤维连接蛋白(K-D=174+/-13 nM)和猪纤毛。F2(271)与100 mU M肝素预孵育可阻断纤毛结合，阻断程度接近69%。用胰酶刮除细胞表面，结合二维液相色谱和串联质谱仪分析，确定Mhp271为表面暴露的。我们的数据表明，Mhp271的R1和R2都参与了与宿主分子的结合。
P>Mycoplasma hyopneumoniae, the causative agent of porcine enzootic pneumonia, adheres to ciliated respiratory epithelia resulting in ciliostasis and epithelial cell death. The cilium adhesin P97 (Mhp183) contains two repeat regions, designated R1 and R2, that play key roles in adherence. Eight pentapeptide repeats in R1 are sufficient to bind porcine cilia; however, both R1 and R2 are needed to bind heparin. Mhp271, a paralogue of P97, is the only other M. hyopneumoniae protein to contain both R1 and R2 repeats. These repeats are arranged as a set of three pentapeptide repeats (designated R1A(271)), two decapeptide repeats (designated R2(271)), and a second set of six pentapeptide repeats (designated R1B(271)). To determine their function, recombinant proteins containing R1A(271) (F1(271)) and R2(271)-R1B(271) (F2(271)) were constructed and used in in vitro binding assays. F2(271), but not F1(271), bound heparin (K-D = 8.1 +/- 0.4 nM), fibronectin (K-D = 174 +/- 13 nM) and porcine cilia. Pre-incubation of F2(271) with 100 mu M heparin blocked cilium binding by similar to 69%. Cell surface shaving with trypsin combined with two-dimensional liquid chromatography coupled to tandem mass spectrometry analysis identified Mhp271 as surface-exposed. Our data suggest that both R1 and R2 in Mhp271 are involved in binding to host molecules.