Abstract
The synthesis of methionine is critical for most bacteria. It is known that cellular methionine has a feedback effect on the expression of met genes involved in de novo methionine biosynthesis. Previous studies revealed that Gram-negative bacteria control met gene expression at the transcriptional level by regulator proteins, while most Gram-positive bacteria regulate met genes at post-transcriptional level by RNA regulators (riboregulators) located in the 5′UTR of met genes. However, despite its importance, the methionine biosynthesis pathway in the Gram-negative Xanthomonas genus that includes many important plant pathogens is completely uncharacterized. Here, we address this issue using the crucifer black rot pathogen Xanthomonas campestris pv. campestris (Xcc), a model bacterium in microbe–plant interaction studies. The work identified an operon (met) involved in de novo methionine biosynthesis in Xcc. Disruption of the operon resulted in defective growth in methionine-limited media and in planta. Western blot analysis revealed that the expression of the operon is dependent on methionine levels. Further molecular analyses demonstrated that the 5′UTR, but not the promoter of the operon, is involved in feedback regulation on operon expression in response to methionine availability, providing an example of a Gram-negative bacterium utilizing a 5′UTR region to control the expression of the genes involved in methionine biosynthesis.
Highlights
To maintain regular protein synthesis, bacterial cells require a constant supply of amino acids
We identified the met operon consisting of three open reading frames (ORFs) that encode enzymes potentially involved in catalysing methionine biosynthesis
The genes encoding the enzymes converting aspartate semialdehyde to cystathionine, i.e. XC1251, XC1252 and XC1253, are located together, while the genes encoding other enzymes involved in the biosynthesis of methionine from aspartate are scattered on the chromosome of Xanthomonas campestris pv. campestris (Xcc) strain 8004 (Fig. 1)
Summary
To maintain regular protein synthesis, bacterial cells require a constant supply of amino acids. Most achieve this by taking up amino acids from the surrounding environment and/ or synthesizing amino acids from simpler compounds (i.e. de novo synthesis). The synthesis of amino acids is considered a complicated and biologically expensive process and is strictly regulated. Of the 20 standard amino acids which are needed for bacterial cell viability, the sulfur-containing methionine [(formyl-)methionine] is somewhat unique. This is because it is the first amino acid at the N-terminus of most proteins and plays an irreplaceable role in the initiation of protein biosynthesis. The methionine derivative S-adenosylmethionine (SAM) serves as a universal methyl group donor in a variety of methyltransferase reactions [2]
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