Everett K D, Hatch T P
Department of Microbiology and Immunology, University of Tennessee, Memphis 38163.
J Bacteriol. 1995 Feb;177(4):877-82. doi: 10.1128/jb.177.4.877-882.1995.
The cysteine-rich envelope proteins of the elementary body form of chlamydiae are thought to be located in the outer membrane on the basis of their insolubility in the weak anionic detergent N-lauryl sarcosinate (Sarkosyl). We found, however, that the insolubility of the small (EnvA) and the large (EnvB) cysteine-rich proteins of Chlamydia psittaci 6BC in Sarkosyl is dependent on the maintenance of a supramolecular disulfide-cross-linked complex and is unlikely to be a valid indicator of outer membrane location. Consequently, we used other methods to characterize the architecture of the cell envelope of C. psittaci 6BC. We found that disulfide-reduced EnvA, previously shown to be a lipoprotein, segregated into the detergent phase during Triton X-114 partitioning experiments and was recovered from the membrane fraction of elementary bodies lysed by nondetergent means. In contrast, disulfide-reduced EnvB segregated to the aqueous phase in partitioning experiments and was found in the soluble fraction of elementary bodies lysed in the absence of detergents. The hydrophobic affinity probe 3-(trifluoromethyl)-3-(m-[125I]iodophenyl)-diazirine labeled the major outer membrane protein and EnvA but did not label EnvB. Treatment of intact elementary bodies of C. psittaci with trypsin had no effect on the cysteine-rich proteins, although the major outer membrane protein was partially degraded. On the basis of these and other observations, we propose that EnvA is anchored to the outer membrane by its lipid moiety, with a hydrophilic peptide portion extending into the periplasm, and that EnvB is located exclusively within the periplasm. We further propose that disulfide-cross-linked polymers of EnvB are the functional equivalent of peptidoglycan, forming a disulfide-cross-linked network with the periplasmic domains of EnvA and other membrane proteins, which accounts for the osmotic stability of elementary bodies.
基于沙眼衣原体原体形式富含半胱氨酸的包膜蛋白在弱阴离子去污剂N-月桂基肌氨酸钠（十二烷基肌氨酸钠）中不溶，推测它们位于外膜中。然而，我们发现鹦鹉热衣原体6BC株的小（EnvA）和大（EnvB）富含半胱氨酸蛋白在十二烷基肌氨酸钠中的不溶性取决于超分子二硫键交联复合物的维持，不太可能是外膜定位的有效指标。因此，我们使用其他方法来表征鹦鹉热衣原体6BC株细胞膜包膜的结构。我们发现，先前显示为脂蛋白的二硫键还原型EnvA在Triton X-114分配实验中分离到去污剂相中，并从非去污剂方法裂解的原体膜部分中回收。相反，二硫键还原型EnvB在分配实验中分离到水相中，并存在于无去污剂裂解的原体可溶部分中。疏水亲和探针3-（三氟甲基）-3-（间-[125I]碘苯基）-二氮杂环丙烷标记了主要外膜蛋白和EnvA，但未标记EnvB。用胰蛋白酶处理完整的鹦鹉热衣原体原体对富含半胱氨酸的蛋白没有影响，尽管主要外膜蛋白部分降解。基于这些和其他观察结果，我们提出EnvA通过其脂质部分锚定在外膜上，亲水肽部分延伸到周质中，并且EnvB仅位于周质内。我们进一步提出，EnvB的二硫键交联聚合物等同于肽聚糖，与EnvA和其他膜蛋白的周质结构域形成二硫键交联网络，这解释了原体的渗透稳定性。
