Localization of BmpA on the exposed outer membrane of Borrelia burgdorferi by monospecific anti-recombinant BmpA rabbit antibodies.
Localization of BmpA on the exposed outer membrane of Borrelia burgdorferi by monospecific anti-recombinant BmpA rabbit antibodies.
复制标题
通过单特异性抗重组 BmpA 兔抗体将 BmpA 定位在伯氏疏螺旋体暴露的外膜上。
DOI：
10.1128/iai.72.4.2280-2287.2004
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发表时间：
2004
影响因子：
3.1
通讯作者：
Cabello,FelipeC
中科院分区：
文献类型：
--
作者：
Shin,JungheeJ;Bryksin,AntonV;Godfrey,HenryP;Cabello,FelipeC
BmpA（P39）是一种免疫显性的染色体编码蛋白。抗BmpA抗体与该旁系同源蛋白家族的其他成员的潜在的强交叉反应性以及先前使用的其对其他Bmp蛋白的反应性未被表征的抗体导致了其在B中的定位的持续争议。burgdorferi。为了提供BmpA定位的明确证明，通过用rBmpB吸收使针对重组BmpA（rBmpA）产生的兔抗体具有单特异性。该试剂与rBmpB、rBmpC或rBmpD在斑点免疫结合中不反应，在B的一维和二维免疫印迹上仅检测到单个39-kDa条带和单个39-kDa，pI 5.0斑点。burgdorferilysates，并从这些裂解物中免疫沉淀单一的39-kDa蛋白。它在B的Triton X-114可溶性和非可溶性组分中检测到BmpA。burgdorferi，表明与细菌细胞内外膜的关联。治疗intactB.用蛋白酶K部分消化BmpA，与细菌外膜上有限的表面暴露一致，这一建议通过未固定B的免疫荧光证实。在体外和体内培养。抗rBmpA抗体对B. burgdorferiB 31在培养物中，再次表明BmpA在暴露的螺旋体外表面上的定位。BmpA的表面定位，抗rBmpA抗体的生长抑制，以及以前报道的不同B. burgdorferisensu lato菌株可能表明BmpA在B. burgdorferibiology.
BmpA (P39) is an immunodominant chromosomally encodedBorrelia burgdorferiprotein. The potential strong cross-reactivity of anti-BmpA antibodies with the other members of this paralogous protein family and the previous use of antibodies whose reactivity to the other Bmp proteins was uncharacterized have resulted in continued controversy over its localization inB. burgdorferi. In an effort to provide a definitive demonstration of the localization of BmpA, rabbit antibodies raised to recombinant BmpA (rBmpA) were rendered monospecific by absorption with rBmpB. This reagent did not react with rBmpB, rBmpC, or rBmpD in dot immunobinding, detected only a single 39-kDa band and a single 39-kDa, pI 5.0 spot on one- and two-dimensional immunoblots ofB. burgdorferilysates, respectively, and immunoprecipitated a single 39-kDa protein from these lysates. It detected BmpA in the Triton X-114-soluble and -insoluble fractions ofB. burgdorferi, suggesting association with both inner and outer bacterial cell membranes. Treatment of intactB. burgdorferiwith proteinase K partially digested BmpA, consistent with a limited surface exposure on the outer bacterial membrane, a suggestion confirmed by immunofluorescence of unfixedB. burgdorfericultured in vitro and in vivo. Anti-rBmpA antibody was bacteriostatic forB. burgdorferiB31 in culture, again suggesting localization of BmpA on the exposed spirochetal outer surface. Surface localization of BmpA, growth inhibition by anti-rBmpA antibodies, and the previously reported conservation ofbmpAin differentB. burgdorferisensu lato strains may indicate that BmpA plays an essential role inB. burgdorferibiology.