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摘要
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.
- 出版日期2014-2-8
- 单位浙江大学
