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The methionine salvage pathway in Bacillus subtilis.

Sekowska A, Danchin A - BMC Microbiol. (2002)

Bottom Line: Among the most remarkable discoveries in this pathway is the role of an analog of ribulose diphosphate carboxylase (Rubisco, the plant enzyme used in the Calvin cycle which recovers carbon dioxide from the atmosphere) as a major step in MTR recycling.In particular, a paralogue or Rubisco, MtnW, is used at one of the steps in the pathway.A major observation is that in the absence of MtnW, MTR becomes extremely toxic to the cell, opening an unexpected target for new antimicrobial drugs.

View Article: PubMed Central - HTML - PubMed

Affiliation: HKU-Pasteur Research Centre, Dexter HC Man Building, 8, Sassoon Road, Pokfulam, Hong Kong, China. sekowska@hkucc.hku.hk

ABSTRACT

Background: Polyamine synthesis produces methylthioadenosine, which has to be disposed of. The cell recycles it into methionine through methylthioribose (MTR). Very little was known about MTR recycling for methionine salvage in Bacillus subtilis.

Results: Using in silico genome analysis and transposon mutagenesis in B. subtilis we have experimentally uncovered the major steps of the dioxygen-dependent methionine salvage pathway, which, although similar to that found in Klebsiella pneumoniae, recruited for its implementation some entirely different proteins. The promoters of the genes have been identified by primer extension, and gene expression was analyzed by Northern blotting and lacZ reporter gene expression. Among the most remarkable discoveries in this pathway is the role of an analog of ribulose diphosphate carboxylase (Rubisco, the plant enzyme used in the Calvin cycle which recovers carbon dioxide from the atmosphere) as a major step in MTR recycling.

Conclusions: A complete methionine salvage pathway exists in B. subtilis. This pathway is chemically similar to that in K. pneumoniae, but recruited different proteins to this purpose. In particular, a paralogue or Rubisco, MtnW, is used at one of the steps in the pathway. A major observation is that in the absence of MtnW, MTR becomes extremely toxic to the cell, opening an unexpected target for new antimicrobial drugs. In addition to methionine salvage, this pathway protects B. subtilis against dioxygen produced by its natural biotope, the surface of leaves (phylloplane).

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Alignment of MtnZ with the consensus of pfam03079 , coding for aci-reductone enzymes. Red letters represent identities, blue letters conservative replacements (classes: AGPST, ILMV, FWY, DENQ, HKR).
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Figure 8: Alignment of MtnZ with the consensus of pfam03079 , coding for aci-reductone enzymes. Red letters represent identities, blue letters conservative replacements (classes: AGPST, ILMV, FWY, DENQ, HKR).

Mentions: The methionine salvage pathway has been deciphered in K. pneumoniae. It is possible, combining this knowledge to the genetic and physiologic results just described, to use it at the basis for reconstructing in silico the corresponding metabolic pathway in B. subtilis. The first steps are similar in both organisms: methylthioadenosine is converted into MTR by a nucleosidase (MtnA, [5]). Subsequently, MTR is phosphorylated into MTR-1-phosphate by MtnK [6]. On the other end of the pathway, methionine is synthesized directly from its keto acid precursor, 2-keto-4-methylthiobutyrate, by a transaminase. MtnV is the likely preferred enzyme for this activity. In K. pneumoniae a dioxygenase is converting 2,3-diketo-5-methylthio-1-phosphopentane into 2-keto-4-methylthiobutyrate [2]. Using dynamic programming (FASTA) we compared the sequence of the corresponding protein to the complete proteome of B. subtilis. YkrZ comes out as the first hit, as the most similar enzyme present in the proteome. Furthermore, it displays a strong consensus similarity with the dioxygenases of the family pfam03079 (Fig. 8) [13]. In order to check whether dioxygen was indeed involved in the case of B. subtilis we grew the cells anaerobically, with nitrate as an electron acceptor, and tested for growth on MTR: while the wild type strain grew well when sulfate was the carbon source, it failed to grow with MTR (Table 1).


The methionine salvage pathway in Bacillus subtilis.

Sekowska A, Danchin A - BMC Microbiol. (2002)

Alignment of MtnZ with the consensus of pfam03079 , coding for aci-reductone enzymes. Red letters represent identities, blue letters conservative replacements (classes: AGPST, ILMV, FWY, DENQ, HKR).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC113757&req=5

Figure 8: Alignment of MtnZ with the consensus of pfam03079 , coding for aci-reductone enzymes. Red letters represent identities, blue letters conservative replacements (classes: AGPST, ILMV, FWY, DENQ, HKR).
Mentions: The methionine salvage pathway has been deciphered in K. pneumoniae. It is possible, combining this knowledge to the genetic and physiologic results just described, to use it at the basis for reconstructing in silico the corresponding metabolic pathway in B. subtilis. The first steps are similar in both organisms: methylthioadenosine is converted into MTR by a nucleosidase (MtnA, [5]). Subsequently, MTR is phosphorylated into MTR-1-phosphate by MtnK [6]. On the other end of the pathway, methionine is synthesized directly from its keto acid precursor, 2-keto-4-methylthiobutyrate, by a transaminase. MtnV is the likely preferred enzyme for this activity. In K. pneumoniae a dioxygenase is converting 2,3-diketo-5-methylthio-1-phosphopentane into 2-keto-4-methylthiobutyrate [2]. Using dynamic programming (FASTA) we compared the sequence of the corresponding protein to the complete proteome of B. subtilis. YkrZ comes out as the first hit, as the most similar enzyme present in the proteome. Furthermore, it displays a strong consensus similarity with the dioxygenases of the family pfam03079 (Fig. 8) [13]. In order to check whether dioxygen was indeed involved in the case of B. subtilis we grew the cells anaerobically, with nitrate as an electron acceptor, and tested for growth on MTR: while the wild type strain grew well when sulfate was the carbon source, it failed to grow with MTR (Table 1).

Bottom Line: Among the most remarkable discoveries in this pathway is the role of an analog of ribulose diphosphate carboxylase (Rubisco, the plant enzyme used in the Calvin cycle which recovers carbon dioxide from the atmosphere) as a major step in MTR recycling.In particular, a paralogue or Rubisco, MtnW, is used at one of the steps in the pathway.A major observation is that in the absence of MtnW, MTR becomes extremely toxic to the cell, opening an unexpected target for new antimicrobial drugs.

View Article: PubMed Central - HTML - PubMed

Affiliation: HKU-Pasteur Research Centre, Dexter HC Man Building, 8, Sassoon Road, Pokfulam, Hong Kong, China. sekowska@hkucc.hku.hk

ABSTRACT

Background: Polyamine synthesis produces methylthioadenosine, which has to be disposed of. The cell recycles it into methionine through methylthioribose (MTR). Very little was known about MTR recycling for methionine salvage in Bacillus subtilis.

Results: Using in silico genome analysis and transposon mutagenesis in B. subtilis we have experimentally uncovered the major steps of the dioxygen-dependent methionine salvage pathway, which, although similar to that found in Klebsiella pneumoniae, recruited for its implementation some entirely different proteins. The promoters of the genes have been identified by primer extension, and gene expression was analyzed by Northern blotting and lacZ reporter gene expression. Among the most remarkable discoveries in this pathway is the role of an analog of ribulose diphosphate carboxylase (Rubisco, the plant enzyme used in the Calvin cycle which recovers carbon dioxide from the atmosphere) as a major step in MTR recycling.

Conclusions: A complete methionine salvage pathway exists in B. subtilis. This pathway is chemically similar to that in K. pneumoniae, but recruited different proteins to this purpose. In particular, a paralogue or Rubisco, MtnW, is used at one of the steps in the pathway. A major observation is that in the absence of MtnW, MTR becomes extremely toxic to the cell, opening an unexpected target for new antimicrobial drugs. In addition to methionine salvage, this pathway protects B. subtilis against dioxygen produced by its natural biotope, the surface of leaves (phylloplane).

Show MeSH
Related in: MedlinePlus