A novel autotransporter of uropathogenic Proteus mirabilis is both a cytotoxin and an agglutinin
A novel autotransporter of uropathogenic Proteus mirabilis is both a cytotoxin and an agglutinin
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DOI：
10.1111/j.1365-2958.2008.06199.x
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发表时间：
2008-05-01
影响因子：
3.6
通讯作者：
Mobley, Harry L. T.
中科院分区：
文献类型：
--
作者：
Alamuri, Praveen;Mobley, Harry L. T.
奇异变形杆菌六种预测的自转运蛋白（ATs）之一，开放阅读框c2341，被预测含有一个丝氨酸蛋白酶基序，并且早些时候被鉴定为奇异变形杆菌中的一种免疫原性外膜蛋白。这个3.2kb的基因编码一个117kDa的蛋白质，它具有一个58个氨基酸长的信号肽、一个75kDa长的N端乘客结构域和一个30kDa长的C端转运结构域。亲和纯化的110kDa自转运蛋白表现出类胰凝乳蛋白酶活性，并且在最适pH值为8.5 - 9.0时以Ca²⁺依赖的方式水解N - 琥珀酰 - 丙氨酸 - 丙氨酸 - 脯氨酸 - 苯丙氨酸 - 对硝基苯胺（N - Suc - Ala - Ala - Pro - Phe - pNa）和N - 琥珀酰 - 丙氨酸 - 丙氨酸 - 脯氨酸 - 亮氨酸 - 对硝基苯胺（N - Suc - Ala - Ala - Pro - Leu - pNa），其Km值分别为22μM和31μM。该活性被枯草杆菌蛋白酶特异性抑制剂亮抑蛋白酶肽和抑糜蛋白酶素抑制。细胞相关形式和纯化形式都对培养的肾脏和膀胱上皮细胞产生细胞病变效应。底物水解以及细胞毒性与乘客结构域相关，并且在任何一个催化残基（丝氨酸366、组氨酸147和天冬氨酸533）发生突变时都会受到影响。在碱性pH值和最适细胞密度下，自转运蛋白还促进奇异变形杆菌的自身聚集，并且这个功能与其蛋白酶活性无关。在奇异变形杆菌的一个同基因pta突变体中，细胞毒性、自身聚集和毒力都显著降低。奇异变形杆菌毒性凝集素（Pta）代表一种新型的自转运细胞毒素，没有细菌同源物，在碱化的尿路中作用最佳，这是奇异变形杆菌感染过程中脲酶介导的尿素水解的一个特征。
One of the six predicted Proteus mirabilis autotransporters (ATs), ORF c2341, is predicted to contain a serine protease motif and was earlier identified as an immunogenic outer membrane protein in P. mirabilis. The 3.2 kb gene encodes a 117 kDa protein with a 58-amino-acid-long signal peptide, a 75-kDa-long N-terminal passenger domain and a 30-kDa-long C-terminal translocator. Affinity-purified 110 kDa AT exhibited chymotrypsin-like activity and hydrolysed N-Suc-Ala-Ala-Pro-Phe-pNa and N-Suc-Ala-Ala-Pro-Leu-pNa with a K-M of 22 mu M and 31 mu M, respectively, under optimal pH of 8.5-9.0 in a Ca2+-dependent manner. Activity was inhibited by subtilase-specific inhibitors leupeptin and chymostatin. Both the cell-associated and purified form elicited cytopathic effects on cultured kidney and bladder epithelial cells. Substrate hydrolysis as well as cytotoxicity was associated with the passenger domain and was compromised upon mutation of any of the catalytic residues (Ser366, His147 and Asp533). At alkaline pH and optimal cell density, the AT also promoted autoaggregation of P. mirabilis and this function was independent of its protease activity. Cytotoxicity, autoaggregation and virulence were significantly reduced in an isogenic pta mutant of P. mirabilis. Proteus toxic agglutinin (Pta) represents a novel autotransported cytotoxin with no bacterial homologues that works optimally in the alkalinized urinary tract, a characteristic of urease-mediated urea hydrolysis during P. mirabilis infection.