Hernández-Allés Santiago, Albertí Sebastián, Álvarez Dolores, Doménech-Sánchez Antonio, Martínez-Martínez Luis, Gil José, Tomás Juan M, Benedí Vicente J
Área de Microbiología, Departamento de Biología, Universidad de las Islas Baleares, Palma de Mallorca, and Unidad de Recursos Naturales, Instituto Mediterráneo de Estudios Avanzados, IMEDEA (CSIC-UIB), Carretera de Valldemosa Km 7·5, E-07071 Palma de Mallorca, Spain.
Departamento de Microbiología, Universidad de Sevilla, Avenida Sánchez Pizjuan s/n, E-41080 Sevilla, Spain.
Microbiology (Reading). 1999 Mar;145 ( Pt 3):673-679. doi: 10.1099/13500872-145-3-673.
Two porins, OmpK36 and OmpK35, have been described previously in Klebsiella pneumoniae, and they are homologous to the Escherichia coli porins OmpC and OmpF, respectively, at both the DNA and amino acid levels. Optimal resolution of the two K. pneumoniae porins by electrophoresis on polyacrylamide gels is not achieved using gel systems already described for E. coli and requires modifications of the bisacrylamide content of the resolving gels. Once resolved, identification of porins OmpK36 and OmpK35 cannot be based solely on their apparent molecular masses since in some strains the OmpK36 porin migrates faster than the OmpK35 porin, whilst in other strains OmpK35 is the faster-migrating porin. Expression of OmpK35 porin is increased in low-osmolarity medium and, combined with Western blot analysis, this allows for the identification of both porins. Application of this identification system showed that most isolates lacking expression of extended-spectrum beta-lactamases express the two porins, whereas most isolates producing these beta-lactamases express only porin OmpK36, and the OmpK35 porin is either very low or not expressed.
此前已在肺炎克雷伯菌中描述了两种孔蛋白，即OmpK36和OmpK35，它们在DNA和氨基酸水平上分别与大肠杆菌孔蛋白OmpC和OmpF同源。使用已描述的用于大肠杆菌的凝胶系统，无法通过聚丙烯酰胺凝胶电泳对肺炎克雷伯菌的这两种孔蛋白进行最佳分辨率分析，需要对分离胶的双丙烯酰胺含量进行调整。一旦分离出来，仅根据孔蛋白OmpK36和OmpK35的表观分子量无法对其进行鉴定，因为在某些菌株中，OmpK36孔蛋白的迁移速度比OmpK35孔蛋白快，而在其他菌株中，OmpK35是迁移速度较快的孔蛋白。OmpK35孔蛋白在低渗培养基中的表达增加，结合蛋白质印迹分析，这有助于鉴定这两种孔蛋白。应用该鉴定系统表明，大多数缺乏超广谱β-内酰胺酶表达的分离株表达这两种孔蛋白，而大多数产生这些β-内酰胺酶的分离株仅表达孔蛋白OmpK36，且OmpK35孔蛋白表达极低或不表达。
