DNA photolyase from the fungus Neurospora crassa. Purification, characterization and comparison with other photolyases - PubMed (original) (raw)

Comparative Study

DNA photolyase from the fungus Neurospora crassa. Purification, characterization and comparison with other photolyases

A P Eker et al. Photochem Photobiol. 1994 Aug.

Abstract

A phr-gene from the filamentous fungus Neurospora crassa was overexpressed in Escherichia coli cells, yielding a biologically active photolyase. After purification till apparent homogeneity, the 66 kDa protein was found to contain equimolar amounts of 5,10-methenyltetrahydrofolic acid (MTHF) and FAD, classifying it as an MTHF-type photolyase. Compared to other MTHF photolyases the absorption maximum of Neurospora photolyase is shifted from ca 380 nm to 391 nm (epsilon = 34,800), while an additional shoulder is present at 465 nm. In dark-adapted enzyme the FAD chromophore is predominantly present in the oxidized form, in contrast with E. coli and Saccharomyces cerevisiae photolyase, which contain mainly semiquinone or fully reduced FAD, respectively. Preillumination or dithionite treatment converted oxidized FAD in Neurospora photolyase into the fully reduced form, with a concomitant shift of the absorption maximum from 391 to 396 nm and disappearance of the 465 nm shoulder. The action spectrum of photoreactivation coincides with the absorption spectrum of preilluminated (reduced) photolyase, extending the spectral region of MTHF-type photolyases from 380 till 396 nm. A quantum yield of 0.57 was obtained for the overall repair reaction. Comparison of spectral properties of FAD in Neurospora photolyase and the model compound lumiflavin points to an apolar microenvironment of photolyase-bound FAD. Neurospora photolyase has distinct advantages over E. coli photolyase as it is more stable and contains a full complement of chromophores.

PubMed Disclaimer

Similar articles

• Cloning and functional characterization of a eucaryotic DNA photolyase gene from Neurospora crassa.
  Yajima H, Inoue H, Oikawa A, Yasui A. Yajima H, et al. Nucleic Acids Res. 1991 Oct 11;19(19):5359-62. doi: 10.1093/nar/19.19.5359. Nucleic Acids Res. 1991. PMID: 1833725 Free PMC article.
• Purification and characterization of Drosophila melanogaster photolyase.
  Kim ST, Malhotra K, Ryo H, Sancar A, Todo T. Kim ST, et al. Mutat Res. 1996 Jun 12;363(2):97-104. doi: 10.1016/0921-8777(96)00003-1. Mutat Res. 1996. PMID: 8676930
• Characterization of a Neurospora crassa photolyase-deficient mutant generated by repeat induced point mutation of the phr gene.
  Shimura M, Ito Y, Ishii C, Yajima H, Linden H, Harashima T, Yasui A, Inoue H. Shimura M, et al. Fungal Genet Biol. 1999 Oct;28(1):12-20. doi: 10.1006/fgbi.1999.1158. Fungal Genet Biol. 1999. PMID: 10512668
• Light-driven enzymatic catalysis of DNA repair: a review of recent biophysical studies on photolyase.
  Weber S. Weber S. Biochim Biophys Acta. 2005 Feb 25;1707(1):1-23. doi: 10.1016/j.bbabio.2004.02.010. Biochim Biophys Acta. 2005. PMID: 15721603 Review.
• Photochemistry, photophysics, and mechanism of pyrimidine dimer repair by DNA photolyase.
  Kim ST, Sancar A. Kim ST, et al. Photochem Photobiol. 1993 May;57(5):895-904. doi: 10.1111/j.1751-1097.1993.tb09232.x. Photochem Photobiol. 1993. PMID: 8337263 Review.

Cited by

• Functional Characterization of Pseudoidium neolycopersici Photolyase Reveals Mechanisms Behind the Efficacy of Nighttime UV on Powdery Mildew Suppression.
  Pathak R, Ergon Å, Stensvand A, Gislerød HR, Solhaug KA, Cadle-Davidson L, Suthaparan A. Pathak R, et al. Front Microbiol. 2020 May 29;11:1091. doi: 10.3389/fmicb.2020.01091. eCollection 2020. Front Microbiol. 2020. PMID: 32547521 Free PMC article.
• Characterization of a cold-adapted DNA photolyase from C. psychrerythraea 34H.
  Munshi S, Rajamoorthi A, Stanley RJ. Munshi S, et al. Extremophiles. 2017 Sep;21(5):919-932. doi: 10.1007/s00792-017-0953-z. Epub 2017 Jul 19. Extremophiles. 2017. PMID: 28726126 Free PMC article.
• The Two Cryptochrome/Photolyase Family Proteins Fulfill Distinct Roles in DNA Photorepair and Regulation of Conidiation in the Gray Mold Fungus Botrytis cinerea.
  Cohrs KC, Schumacher J. Cohrs KC, et al. Appl Environ Microbiol. 2017 Aug 17;83(17):e00812-17. doi: 10.1128/AEM.00812-17. Print 2017 Sep 1. Appl Environ Microbiol. 2017. PMID: 28667107 Free PMC article.
• A photolyase-like protein from Agrobacterium tumefaciens with an iron-sulfur cluster.
  Oberpichler I, Pierik AJ, Wesslowski J, Pokorny R, Rosen R, Vugman M, Zhang F, Neubauer O, Ron EZ, Batschauer A, Lamparter T. Oberpichler I, et al. PLoS One. 2011;6(10):e26775. doi: 10.1371/journal.pone.0026775. Epub 2011 Oct 31. PLoS One. 2011. PMID: 22066008 Free PMC article.
• Neurospora crassa Light Signal Transduction Is Affected by ROS.
  Belozerskaya TA, Gessler NN, Isakova EP, Deryabina YI. Belozerskaya TA, et al. J Signal Transduct. 2012;2012:791963. doi: 10.1155/2012/791963. Epub 2011 Oct 20. J Signal Transduct. 2012. PMID: 22046507 Free PMC article.

Publication types

MeSH terms

Substances

LinkOut - more resources

• Full Text Sources

  • Wiley