DOMAIN-STRUCTURE AND RNA ANNEALING ACTIVITY OF THE ESCHERICHIA-COLI REGULATORY PROTEIN STPA

The Escherichia coil regulatory protein StpA bears striking similarity to the chromatin-associated protein H-NS. These two proteins have man y structural, functional and mechanistic parallels. Although H-NS is m ore abundant in the cell, both proteins act as transcriptional regulat ors, both bind to curved DNA and both restrain DNA supercoils. However , StpA is better able to promote RNA annealing and trans-splicing in v itro. In this study, phylogenetic analyses and experiments to examine the protease sensitivity of StpA and H-NS suggest a similar structure for the two proteins, Both proteins consist of two structured domains separated by an exposed protease-sensitive linker, The N-terminal (Stp A-NterL) and C-terminal (StpA-CterL) domains of StpA, as well as the f ull-length StpA and H-NS proteins, were cloned, overproduced in E. coi l and purified to homogeneity, StpA-CterL, but not StpA-NterL, promote s strand annealing of complementary RNA oligonucleotides and in vitro trans-splicing of a model group I intron. Both StpA and StpA-CterL exh ibited stronger RNA-modulating activity than H-NS, Phylogenetic analys es showed that the N-terminal and C-terminal domains can exist autonom ously. The phylogenetic and experimental data are compatible with a tw o-domain model for StpA and H-NS, with independently functioning modul es joined by a non-conserved linker, and with the observed RNA-related activities residing entirely within the C-terminal domain.