Two Glycine Riboswitches Activate the Glycine Cleavage System Essential for Glycine Detoxification in Streptomyces griseus

Abstract

The glycine cleavage (GCV) system catalyzes the oxidative cleavage of glycine into CO 2 , NH 4 ؉ , and a methylene group, which is accepted by tetrahydrofolate (THF) to form N 5 ,N 10 -methylene-THF. Streptomyces griseus contains gcvP and the gcvT-gcvH operon, which encode three intrinsic components of the GCV system. We identified the transcriptional start sites of gcvTH and gcvP and found putative glycine riboswitches in their 5= untranslated regions (5= UTRs). The ratios of the transcripts of the gcvT and gcvP coding sequences (CDSs) to those of the respective 5= UTRs were significantly higher in the presence of glycine in the wild-type strain. However, the levels of gcvT and gcvP CDS transcripts were not increased by glycine in the respective 5= UTR deletion mutants. A reporter gene assay showed that a transcriptional terminator exists in the 5= UTR of gcvTH. Furthermore, by an in-line probing assay, we confirmed that glycine bound directly to the putative riboswitch RNAs. These results indicate that the S. griseus glycine riboswitches enhance transcriptional read-through to the downstream CDSs, like known glycine riboswitches in other bacteria. We examined the growth of three mutants in which either or both of the gcvTH and gcvP 5= UTRs were deleted. Like the wild-type strain, all mutants grew vigorously in a medium containing 0.9% glucose as a carbon source. However, the mutants showed severely restricted growth in a medium containing 0.9% glucose and 1% glycine, while the wildtype strain grew normally. This indicates that glycine has a growth-inhibitory effect and that the GCV system plays a critical role in glycine detoxification in S. griseus.

Glyphosate pathways to modern diseases V: Amino acid analogue of glycine in diverse proteins

“…Glycine is both essential and toxic to bacteria. It is well known that glycine inhibits bacterial growth [233][234][235][236], by substituting for alanine into peptidoglycan precursors [237][238][239]. Glycine-containing precursors are poor substrates for peptidoglycan biosynthesis enzymes as well as for the transpeptidation reaction, leading to both a deficiency in muropeptides and a high percentage of muropeptides that are not cross-linked.…”

“…As a consequence of glycine's toxicity, it is important for bacteria to be able to quickly break glycine down into basic building blocks. Oxidative cleavage to CO 2 , + 4 NH and a methyl group is carried out by the glycine cleavage system (GCS), and the methyl group becomes a major source for one-carbon metabolism, beginning with the conversion of tetrahydrofolate (THF) to methylene-THF [237], which is then used to biosynthesize various cellular compounds, including, importantly, purines and methionine. The GCS also produces NADH in the oxidative cleavage step, which yields energy through the electron transport system.…”

“…In this way, glycine is quickly cleared and put to good use, fueling the electron transport chain and the one-carbon metabolism pathways. Glycine is highly toxic to mutants missing these riboswitch regions; a medium containing only 1% glycine severely restricts their growth [237].…”

Singlet glycine riboswitches bind ligand as well as tandem riboswitches

“…Glycine riboswitches control genes related to the import, synthesis, and degradation of glycine, and these genes maintain the intracellular glycine concentration within a narrow range in bacterial cells (Barrick and Breaker 2007;Barrick et al 2004;Kazanov et al 2007). Disruption of the riboswitches has been shown to be toxic to S. griseus growing in media containing glycine (Tezuka and Ohnishi 2014). More than 7000 examples of the riboswitch class are found across bacterial phyla and environmental sequences (P McCown and R Breaker, unpubl.…”

Ligand binding by the tandem glycine riboswitch depends on aptamer dimerization but not double ligand occupancy

“…In addition to exploiting excess glycine as an energy source, bacteria must regulate concentrations to prevent glycine toxicity. High concentrations of glycine interfere with cell wall biosynthesis, and glycine riboswitches are necessary for Streptomyces griseus populations to grow in high glycine (Tezuka and Ohnishi 2014). The riboswitch system allows organisms to respond quickly to changes in local environment, catabolizing glycine for energy when environmental concentrations get too high, and importing glycine from the environment when concentrations fall too low.…”