Pyridoxal-5′-phosphate as an oxygenase cofactor: Discovery of a carboxamide-forming, α-amino acid monooxygenase-decarboxylase

Ying Huang, Xiaodong Liu, Zheng Cui, Daniel Wiegmann, Giuliana Niro, Christian Ducho, Yuan Song, Zhaoyong Yang, Steven G. Van Lanen

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

Capuramycins are antimycobacterial antibiotics that consist of a modified nucleoside named uridine-5′-carboxamide (CarU). Previous biochemical studies have revealed that CarU is derived from UMP, which is first converted to uridine-5′-aldehyde in a reaction catalyzed by the dioxygenase CapA and subsequently to 5′-C-glycyluridine (GlyU), an unusual β–hydroxy-α-amino acid, in a reaction catalyzed by the pyridoxal-5′-phosphate (PLP)-dependent transaldolase CapH. The remaining steps that are necessary to furnish CarU include decarboxylation, O atom insertion, and oxidation. We demonstrate that Cap15, which has sequence similarity to proteins annotated as bacterial, PLP-dependent L-seryl-tRNA (Sec) selenium transferases, is the sole catalyst responsible for complete conversion of GlyU to CarU. Using a complementary panel of in vitro assays, Cap15 is shown to be dependent upon substrates O2 and (5′S,6′R)-GlyU, the latter of which was unexpected given that (5′S,6′S)-GlyU is the isomeric product of the transaldolase CapH. The two products of Cap15 are identified as the carboxamide-containing CarU and CO2. While known enzymes that catalyze this type of chemistry, namely α-amino acid 2-mono-oxygenase, utilize flavin adenine dinucleotide as the redox cofactor, Cap15 remarkably requires only PLP. Furthermore, Cap15 does not produce hydrogen peroxide and is shown to directly incorporate a single O atom from O2 into the product CarU and thus is an authentic PLP-dependent monooxygenase. In addition to these unusual discoveries, Cap15 activity is revealed to be dependent upon the inclusion of phosphate. The biochemical characteristics along with initiatory mechanistic studies of Cap15 are reported, which has allowed us to assign Cap15 as a PLP-dependent (5′S,6′R)-GlyU: O2 monooxygenase-decarboxylase.

Original languageEnglish
Pages (from-to)974-979
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume115
Issue number5
DOIs
StatePublished - Jan 30 2018

Bibliographical note

Funding Information:
ACKNOWLEDGMENTS. We thank Dr. Koichi Nonaka (Daiichi-Sankyo Co.) for intellectual contributions and for providing strains and cosmids. This work was supported in part by National Institutes of Health Grants AI128862 and AI087849 (to S.G.V.L.).

Keywords

  • Antibiotic
  • Biosynthesis
  • Enzyme function
  • Natural products

ASJC Scopus subject areas

  • General

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