Fan F, Macnab R M
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520-8114, USA.
J Biol Chem. 1996 Dec 13;271(50):31981-8. doi: 10.1074/jbc.271.50.31981.
FliI is a protein needed for flagellar assembly in Salmonella typhimurium. It shows sequence similarity to the catalytic beta subunit of the F0F1-ATPase and is even more closely related to putative ATPases in Type III bacterial secretory pathways. A His-tagged version of FliI, which was fully functional in complementation tests, was purified to homogeneity. It had an ATPase activity of 0.16 s-1 at 25 degrees C and pH 7, and a Km for ATP of 0.3 mM; Mg2+ was required. The activity was not affected by inhibitors of the F-, V- or P-type ATPases, or inhibitors of the Type I or Type II bacterial secretory pathways. Mutations K188I and Y363S decreased the ATPase activity about 100-fold, increased the Km about 10-fold, blocked flagellar assembly, and were dominant. Other FliI mutations that disrupted flagellar protein export were found near the N terminus; they permitted essentially wild-type ATPase activity, were not dominant, and showed a dosage-dependent phenotype. We propose that FliI has a C-terminal ATPase domain and an N-terminal domain that interacts with other components in the flagellum-specific export apparatus.
FliI是鼠伤寒沙门氏菌鞭毛组装所需的一种蛋白质。它与F0F1 - ATP酶的催化β亚基具有序列相似性，并且与III型细菌分泌途径中的假定ATP酶关系更为密切。一种在互补试验中具有完全功能的His标签FliI版本被纯化至同质。它在25℃和pH 7时的ATP酶活性为0.16 s-1，ATP的Km为0.3 mM；需要Mg2+。该活性不受F型、V型或P型ATP酶抑制剂或I型或II型细菌分泌途径抑制剂的影响。K188I和Y363S突变使ATP酶活性降低约100倍，Km增加约10倍，阻断鞭毛组装，且具有显性。在N端附近发现了其他破坏鞭毛蛋白输出的FliI突变；它们具有基本野生型的ATP酶活性，不具有显性，并且表现出剂量依赖性表型。我们提出FliI具有一个C端ATP酶结构域和一个N端结构域，该N端结构域与鞭毛特异性输出装置中的其他成分相互作用。
