Effects of Tryptophan Starvation on Levels of the trp RNA-Binding Attenuation Protein (TRAP) and Anti-TRAP Regulatory Protein and Their Influence on trp Operon Expression in Bacillus subtilis

Abstract

The anti-TRAP protein (AT), encoded by the rtpA gene of Bacillus subtilis, can bind to and inhibit the tryptophan-activated trp RNA-binding attenuation protein (TRAP). AT binding can prevent TRAP from promoting transcription termination in the leader region of the trp operon, thereby increasing trp operon expression. We show here that AT levels continue to increase as tryptophan starvation becomes more severe, whereas the TRAP level remains relatively constant and independent of tryptophan starvation. Assuming that the functional form of AT is a trimer, we estimate that the ratios of AT trimers per TRAP molecule are 0.39 when the cells are grown under mild tryptophan starvation conditions, 0.83 under more severe starvation conditions, and approximately 2.0 when AT is expressed maximally. As the AT level is increased, a corresponding increase is observed in the anthranilate synthase level. When AT is expressed maximally, the anthranilate synthase level is about 70% of the level observed in a strain lacking TRAP. In a nutritional shift experiment where excess phenylalanine and tyrosine could potentially starve cells of tryptophan, both the AT level and anthranilate synthase activity were observed to increase. Expression of the trp operon is clearly influenced by the level of AT.