New function for Escherichia coli xanthosine phophorylase (xapA): genetic and biochemical evidences on its participation in NAD+ salvage from nicotinamide
| Autor: | Li-Xin Xiang, Wei-ren Dong, Jian-Zhong Shao, Cen-Cen Sun, Guan-qun Zhu, Shi-Hua Hu |
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| Rok vydání: | 2014 |
| Předmět: |
Niacinamide
Microbiology (medical) Purine nucleoside phosphorylase Biology medicine.disease_cause Microbiology Gene Knockout Techniques chemistry.chemical_compound Pyridine nucleotide cycles Escherichia coli medicine Pentosyltransferases Salvage pathway Nucleotide salvage Nicotinamide Escherichia coli Proteins NAD salvage Xanthosine NAD Nicotinamide riboside Culture Media Purine-Nucleoside Phosphorylase chemistry Biochemistry NAD+ kinase Research Article |
| Zdroj: | BMC Microbiology |
| ISSN: | 1471-2180 |
| Popis: | Background In an effort to reconstitute the NAD+ synthetic pathway in Escherichia coli (E. coli), we produced a set of gene knockout mutants with deficiencies in previously well-defined NAD+de novo and salvage pathways. Unexpectedly, the mutant deficient in NAD+de novo and salvage pathway I could grow in M9/nicotinamide medium, which was contradictory to the proposed classic NAD+ metabolism of E. coli. Such E. coli mutagenesis assay suggested the presence of an undefined machinery to feed nicotinamide into the NAD+ biosynthesis. We wanted to verify whether xanthosine phophorylase (xapA) contributed to a new NAD+ salvage pathway from nicotinamide. Results Additional knockout of xapA further slowed down the bacterial growth in M9/nicotinamide medium, whereas the complementation of xapA restored the growth phenotype. To further validate the new function of xapA, we cloned and expressed E. coli xapA as a recombinant soluble protein. Biochemical assay confirmed that xapA was capable of using nicotinamide as a substrate for nicotinamide riboside formation. Conclusions Both the genetic and biochemical evidences indicated that xapA could convert nicotinamide to nicotinamide riboside in E. coli, albeit with relatively weak activity, indicating that xapA may contribute to a second NAD+ salvage pathway from nicotinamide. We speculate that this xapA-mediated NAD+ salvage pathway might be significant in some bacteria lacking NAD+de novo and NAD+ salvage pathway I or II, to not only use nicotinamide riboside, but also nicotinamide as precursors to synthesize NAD+. However, this speculation needs to be experimentally tested. |
| Databáze: | OpenAIRE |
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