INVOLVEMENT OF BOXA NUCLEOTIDES IN THE FORMATION OF A STABLE RIBONUCLEOPROTEIN COMPLEX CONTAINING THE BACTERIOPHAGE-LAMBDA-N PROTEIN

The association of the transcriptional antitermination protein N of ba cteriophage lambda with Escherichia coli RNA polymerase depends on nut site RNA (boxA + boxB) in the nascent transcript and the host protein , NusA. This ribonucleoprotein complex can transcribe through Rho depe ndent and intrinsic termination sites located up to several hundred ba se pairs downstream of nut. For antitermination to occur farther downs tream, this core antitermination complex must be stabilized by the hos t proteins NusB, NusG, and ribosomal protein S10. Here, we show that t he assembly of NusB, NusG, and S10 onto the core complex involves nucl eotides 2-7 of lambda boxA (CGCUCUUACACA) and is a fully cooperative p rocess that depends on the presence of all three proteins. This assemb ly of NusB, NusG, and S10 also requires the carboxyl-terminal region ( amino acids 73-107) of N, which interacts directly with RNA polymerase . NusB and S10 assemble in the absence of NusG when lambda boxA is alt ered at nucleotides 8 and 9 to create a consensus version of boxA (CGC UCUUUAACA). These experiments suggest that multiple protein-protein an d protein-RNA interactions are required to convert a core antiterminat ion complex into a complete complex.