Nicotinamide Adenine Dinucleotides Arrest Photoreduction of Class II DNA Photolyases in FADH ˙ State

Photochem Photobiol. 2018 Jan;94(1):81-87. doi: 10.1111/php.12834. Epub 2017 Oct 26.

Abstract

All light-sensitive members of the photolyase/cryptochrome family rely on FAD as catalytic cofactor. Its activity is regulated by photoreduction, a light-triggered electron transfer process from a conserved tryptophan triad to the flavin. The stability of the reduced flavin depends on available external electron donors and oxygen. In this study, we show for the class II photolyase of Methanosarcina mazei, MmCPDII, that it utilizes physiologically relevant redox cofactors NADH and NADPH for the formation of the semiquinoid FAD in a light-dependent reaction. Using redox-inert variants MmCPDII/W388F and MmCPDII/W360F, we demonstrate that photoreduction by NADH and NADPH requires the class II-specific tryptophan cascade of MmCPDII. Finally, we confirmed that mutations in the tryptophan cascade can be introduced without any substantial structural disturbances by analyzing crystal structures of MmCPDII/W388F, MmCPDII/W360F and MmCPDII/Y345F.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Archaeal Proteins / chemistry
  • Archaeal Proteins / genetics
  • Archaeal Proteins / metabolism*
  • Deoxyribodipyrimidine Photo-Lyase / chemistry
  • Deoxyribodipyrimidine Photo-Lyase / genetics
  • Deoxyribodipyrimidine Photo-Lyase / metabolism*
  • Electron Transport
  • Flavin-Adenine Dinucleotide / metabolism*
  • Kinetics
  • Light
  • Methanosarcina / enzymology*
  • Molecular Conformation
  • Mutation
  • NAD / metabolism*
  • NADP / metabolism
  • Oxidation-Reduction
  • Tryptophan / metabolism

Substances

  • Archaeal Proteins
  • NAD
  • Flavin-Adenine Dinucleotide
  • NADP
  • Tryptophan
  • Deoxyribodipyrimidine Photo-Lyase