Hildebrand M, Aldridge P, Geider K
Max-Planck-Institut für Zellbiologie, Ladenburg, Germany.
Mol Genet Genomics. 2006 Mar;275(3):310-9. doi: 10.1007/s00438-005-0085-5. Epub 2006 Jan 11.
The small basic histone-like protein H-NS is known for bacteria to attenuate virulence of several animal pathogens. An hns homologue from E. amylovora was identified by complementing an E. coli hns-mutant strain with a cosmid library from E. amylovora. A 1.6 kb EcoRI-fragment complemented the mucoid phenotype and repressed the ss-glucosidase activity of E. coli PD32. The open reading frame encoding an H-NS-like protein of 134 amino acid was later shown to be located on plasmid pEA29 (McGhee and Jones 2000). A chromosomal hns gene was amplified with PCR consensus primers and localized near galU of E. amylovora. E. amylovora mutants were created by insertion of a resistance cassette, and the intact gene was inserted into a high copy number plasmid for constitutive expression. Purified chromosomal H-NS protein preferentially bound to a DNA fragment from the lsc region and bending was predicted for an adjacent fragment with the rlsB-promoter. Levan production was significantly increased by hns mutations. Synthesis of the capsular exopolysaccharide amylovoran and of levan were reduced, when hns from the E. amylovora plasmid was overexpressed. A mutation in chromosomal hns of E. amylovora increased amylovoran synthesis, and both mutations retarded symptom formation on immature pears.
小的碱性类组蛋白H-NS因能减弱几种动物病原体的毒力而在细菌中广为人知。通过用来自梨火疫病菌（E. amylovora）的黏粒文库对大肠杆菌hns突变株进行互补，鉴定出了梨火疫病菌的一个hns同源物。一个1.6 kb的EcoRI片段互补了黏液样表型并抑制了大肠杆菌PD32的β-葡萄糖苷酶活性。后来发现，编码一个134个氨基酸的类H-NS蛋白的开放阅读框位于质粒pEA29上（麦吉和琼斯，2000年）。用PCR通用引物扩增了梨火疫病菌的染色体hns基因，并将其定位在梨火疫病菌galU附近。通过插入抗性盒构建了梨火疫病菌突变体，并将完整基因插入高拷贝数质粒中进行组成型表达。纯化的染色体H-NS蛋白优先结合来自lsc区域的一个DNA片段，并且预测与rlsB启动子相邻的一个片段会发生弯曲。hns突变显著增加了果聚糖的产生。当梨火疫病菌质粒中的hns过表达时，荚膜胞外多糖淀粉糊精和果聚糖的合成减少。梨火疫病菌染色体hns的一个突变增加了淀粉糊精的合成，并且这两种突变都延缓了未成熟梨上症状的形成。
