Mutagenesis studies on TenA: A thiamin salvage enzyme from Bacillus subtilis

Jenkins, Amy; Zhang, Yang; Ealick, S.E.; Begley, Tadhg P.

doi:10.1016/j.bioorg.2007.10.005

Cited by 21 publications

(34 citation statements)

References 13 publications

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“…TenA was shown to have 100 times greater activity against aminopyrimidine than with thiamin, further demonstrating that thiamin is not the natural substrate for this enzyme (76). The putative transporter ThiY was also shown to bind aminopyrimidine, which is then delivered to the ThiXZ transport channel.…”

Section: Thiamin Degradationmentioning

confidence: 99%

The Structural and Biochemical Foundations of Thiamin Biosynthesis

Jurgenson

Begley

Ealick

2009

Annu. Rev. Biochem.

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279

323

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Thiamin is synthesized by most prokaryotes and by eukaryotes such as yeast and plants. In all cases, the thiazole and pyrimidine moieties are synthesized in separate branches of the pathway and coupled to form thiamin phosphate. A final phosphorylation gives thiamin pyrophosphate, the active form of the cofactor. Over the past decade or so, biochemical and structural studies have elucidated most of the details of the thiamin biosynthetic pathway in bacteria. Formation of the thiazole requires six gene products, and formation of the pyrimidine requires two. In contrast, details of the thiamin biosynthetic pathway in yeast are only just beginning to emerge. Only one gene product is required for the biosynthesis of the thiazole and one for the biosynthesis of the pyrimidine. Thiamin can also be transported into the cell and can be salvaged through several routes. In addition, two thiamin degrading enzymes have been characterized, one of which is linked to a novel salvage pathway.

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Section: Thiamin Degradationmentioning

confidence: 99%

The Structural and Biochemical Foundations of Thiamin Biosynthesis

Jurgenson

Begley

Ealick

2009

Annu. Rev. Biochem.

Self Cite

279

323

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“…Hydrolytic activity towards the substrate 4‐amino‐5‐aminomethyl‐2‐methylpyrimidine (Interchim, Montluçon, France) was determined, as described previously [17], by monitoring the release of ammonia through the glutamate dehydrogenase assay [24]. Recombinant HP1287 with a concentration of 2.4 μ m , was added to a mixture of 5 units of glutamate dehydrogenase, 5 m m α‐ketoglutarate, 0.1 m m EDTA, 0.250 m m NADPH and 20–480 μ m 4‐amino‐5‐aminomethyl‐2‐methylpyrimidine in two different buffers (20 m m sodium phosphate at pH 8 and 50 m m Mes at pH 6).…”

Section: Methodsmentioning

confidence: 99%

Structural and mutational analysis of TenA protein (HP1287) from the Helicobacter pylori thiamin salvage pathway – evidence of a different substrate specificity

et al. 2009

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HP1287 (tenA) from Helicobacter pylori is included among the genes that play a relevant role in bacterium colonization and persistence. The gene has been cloned and its product, protein TenA, has been expressed and purified. The crystal structures of the wild‐type protein and the mutant F47Y have been determined at resolutions of 2.7 and 2.4 Å, respectively. The molecular model, a homotetramer with 222 symmetry, shows that the H. pylori TenA structure belongs to the thiaminase II class of proteins. These enzymes were recently found to be involved in a salvage pathway for the synthesis of the thiamin precursor hydroxypyrimidine, which constitutes a building block in thiamin biosynthesis, in particular in bacteria living in the soil. By contrast, enzymatic measurements on TenA from H. pylori indicate that the activity on the putative substrate 4‐amino‐5‐aminomethyl‐2‐methylpyrimidine is very modest. Moreover, in the present study, we demonstrate that the mutation at residue 47, a position where a phenylalanine occurs in all the strains of H. pylori sequenced to date, is not sufficient to explain the very low catalytic activity toward the expected substrate. As a result of differences in the colonization environment of H. pylori as well as the TenA structural and catalytic peculiar features, we suggest a possible pivotal role for the H. pylori enzyme in the thiamin biosynthetic route, which is in agreement with the relevance of this protein in the stomach colonization process. Structured digital abstract http://mint.bio.uniroma2.it/mint/search/interaction.do?interactionAc=MINT-7260232: TenA (uniprotkb:http://www.ebi.uniprot.org/entry/O25874) and TenA (uniprotkb:http://www.ebi.uniprot.org/entry/O25874) bind (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0407) by x‐ray crystallography (http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0114)

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“…TenA has been structurally and mechanistically characterized. 58,59 It is mechanistically similar to thiaminase I 60 and catalyzes the substitution reaction by an addition-elimination mechanism. TenA is widely distributed in all three kingdoms of life, suggesting that this salvage pathway is also widely used.…”

Section: Thiamin Salvagementioning

confidence: 99%