Probing the structure of Nun transcription arrest factor bound to RNA polymerase.

The coliphage HK022 protein Nun transcription elongation arrest factor inhibits RNA polymerase translocation. In vivo, Nun acts specifically to block transcription of the coliphage λ chromosome. Using in vitro assays, we demonstrate that Nun cross-links RNA in an RNA:DNA hybrid within a ternary elongation complex (TEC).

Bacteriophage HK022 Nun protein arrests transcription by blocking lateral mobility of RNA polymerase during transcription elongation.

Coliphage HK022 excludes phage λ by subverting the λ antitermination system and arresting transcription on the λ chromosome. The 12 kDa HK022 Nun protein binds to λ nascent transcript through its N-terminal Arginine Rich Motif (ARM), blocking access by λ N and arresting transcription via a C-terminal interaction with RNA polymerase. In a purified in vitro system, we recently demonstrated that Nun arrests transcription by restricting lateral movement of transcription elongation complex (TEC) along the DNA register, thereby freezing the translocation state.

Coliphage HK022 Nun protein inhibits RNA polymerase translocation.

The Nun protein of coliphage HK022 arrests RNA polymerase (RNAP) in vivo and in vitro at pause sites distal to phage λ N-Utilization (nut) site RNA sequences. We tested the activity of Nun on ternary elongation complexes (TECs) assembled with templates lacking the λ nut sequence. We report that Nun stabilizes both translocation states of RNAP by restricting lateral movement of TEC along the DNA register.

An amino acid substitution in a capsid protein enhances phage survival in mouse circulatory system more than a 1000-fold.

In experiments with germ free mice, free from adaptive antibodies to the bacterial virus lambda phage, titers of the virus in the circulatory system have been reported to decrease by more than 10(9)pfu within 48 h of intraperitoneal intravenous or oral administration. Based on these observations, serial passage techniques have been used to select lambda phage mutants, with 13,000-16,000-fold greater capacity to remain in the mouse circulatory system 24h after intraperitoneal injection. In these prior studies the "long-circulating" phage, designated lambdaArgo phage, had at least three mutations including one in the major phage capsid (E) protein, which resulted in the change of glutamic acid to a lysine at residue 158.