S. Shiota et H. Nakayama, UV ENDONUCLEASE OF MICROCOCCUS-LUTEUS, A CYCLOBUTANE PYRIMIDINE DIMER-DNA GLYCOSYLASE ABASIC LYASE - CLONING AND CHARACTERIZATION OF THE GENE/, Proceedings of the National Academy of Sciences of the United Statesof America, 94(2), 1997, pp. 593-598
The gene of Micrococcus luteus UV endonuclease (cyclobutane pyrimidine
dimer-DNA glycosylase/abasic lyase) was cloned and characterized. The
cloned gene, whose product had a predicted molecular mass of 17,120 D
a, was found to be capable of complementing the Escherichia coli uvrA6
mutation in vivo with respect to resistance to acetone-mediated molec
ular photosensitization, a treatment producing exclusively cyclobutane
pyrimidine dimers in DNA. It also generated a nicking activity specif
ic for photosensitization-treated DNA by in vitro transcription/transl
ation. When expressed in E. coil cells, the gene produced a protein st
ructurally identical with UV endonuclease and possessing an activity c
onsistent with cyclobutane pyrimidine dimer-DNA glycosylase/abasic lya
se with respect to the effect of inhibitors and the site of the DNA ba
ckbone scission. Furthermore, the UV endonuclease-deficient mutant DB7
was shown to regain the enzyme through transformation with the cloned
gene. The deduced amino acid sequence of the gene product was at best
27% identical with that of endonuclease V of phage T4, an enzyme stri
kingly similar to UV endonuclease in molecular and catalytic propertie
s. Despite this marginal overall similarity in amino acid sequence, fo
ur of the seven amino acid residues reported to be functionally import
ant in the T4 enzyme were found to be conserved in the M. luteus enzym
e. We propose that the gene be called uveA.