Jul 27, 2001

The subunit structure and catalytic mechanism of the Bacillus subtilis DNA repair enzyme spore photoproduct lyase

Proceedings of the National Academy of Sciences of the United States of America
Roberto Rebeil, Wayne L Nicholson

Abstract

The major DNA photoproduct of dormant, UV-irradiated Bacillus subtilis spores is the thymine dimer 5-thyminyl-5,6-dihydrothymine [spore photoproduct (SP)]. During spore germination, SP is reversed to two intact thymines in situ by the DNA repair enzyme SP lyase, an S-adenosylmethionine (S-AdoMet)-dependent iron-sulfur ([Fe-S]) protein encoded by the splB gene. In the present work, cross-linking, SDS/PAGE, and size exclusion chromatography revealed that SplB protein dimerized when incubated with iron and sulfide under anaerobic reducing conditions. SplB isolated under aerobic conditions generated an EPR spectrum consistent with that of a partially degraded [3Fe-4S] center, and reduction of SplB with dithionite shifted the spectrum to that of a [4Fe-4S] center. Addition of S-AdoMet to SplB converted some of the [4Fe-4S] centers to an EPR-silent form consistent with electron donation to S-AdoMet. HPLC and electrospray ionization MS analyses showed that SP lyase cleaved S-AdoMet to generate 5'-deoxyadenosine. The results indicate that (i) SP lyase is a homodimer of SplB; (ii) dimer formation is coordinated by a [4Fe-4S] center; and (iii) the reduced [4Fe-4S] center is capable of donating electrons to S-AdoMet to generate a 5'-adeno...Continue Reading

  • References24
  • Citations54

References

  • References24
  • Citations54

Citations

Mentioned in this Paper

SGPL1
SDS-PAGE
Base Excision Repair
Protein Conformation
Catalysis
Natto Bacteria
Dimerization
SPL1 protein, Arabidopsis
DNA Photolyase
SOD1

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