TY - JOUR
T1 - phrA, the major photoreactivating factor in the cyanobacterium Synechocystis sp. strain PCC 6803 codes for a cyclobutane-pyrimidine-dimer- specific DNA photolyase
AU - Ng, Wing On
AU - Zentella, Rodolfo
AU - Wang, Yinsheng
AU - Taylor, John Stephen A.
AU - Pakrasi, Himadri B.
N1 - Funding Information:
Acknowledgements RSF1010 and pRL443 are gifts from Dr. Michael Bagdasarian (Department of Microbiology, Michigan State University, East Lansing, Mich.) and Peter Wolk and Jeffery Elhai (DOE Laboratory, Michigan State University, East Lansing, Mich.) respectively. We thank Liping Sun and Wenjin Wang (Chemistry Department, Washington University) for their help with the DNA photolyase assay. Mass spectrometry was provided by the Washington University Mass Spectrometry Resource, an NIH Research Resource (grant no. P41RR0954). This project is supported by the Human Frontier Science Program to H.B.P. Partial support for W.O.N. is provided by a training grant to the Department of Biology from Monsanto Company.
PY - 2000/6/1
Y1 - 2000/6/1
N2 - A new broad-host-range plasmid, pSL1211, was constructed for the over- expression of genes in Synechocystis sp. strain PCC 6803. The plasmid was derived from RSF1010 and an Escherichia coli over-expression plasmid, pTrcHisC. Over-expressed protein is made with a removable N-terminal histidine tag. The plasmid was used to over-express the phrA gene and purify the gene product from Synechocystis sp. strain PCC 6803. PhrA is the major ultraviolet-light-resistant factor in the cyanobacterium. The purified PhrA protein exhibited an optical absorption spectrum similar to that of the cyclobutane pyrimidine dimer (CPD) DNA photolyase from Synechococcus sp. strain PCC 6301 (Anacystis nidulans). Mass spectrometry analysis of PhrA indicated that the protein contains 8-hydroxy-5-deazariboflavin and flavin adenine dinucleotide (FADH2) as cofactors. PhrA repairs only cyclobutane pyrimidine dimer but not pyrimidine (6-4) pyrimidinone photoproducts. On the basis of these results, the PhrA protein is classified as a class I, HDF- type, CPD DNA photolyase.
AB - A new broad-host-range plasmid, pSL1211, was constructed for the over- expression of genes in Synechocystis sp. strain PCC 6803. The plasmid was derived from RSF1010 and an Escherichia coli over-expression plasmid, pTrcHisC. Over-expressed protein is made with a removable N-terminal histidine tag. The plasmid was used to over-express the phrA gene and purify the gene product from Synechocystis sp. strain PCC 6803. PhrA is the major ultraviolet-light-resistant factor in the cyanobacterium. The purified PhrA protein exhibited an optical absorption spectrum similar to that of the cyclobutane pyrimidine dimer (CPD) DNA photolyase from Synechococcus sp. strain PCC 6301 (Anacystis nidulans). Mass spectrometry analysis of PhrA indicated that the protein contains 8-hydroxy-5-deazariboflavin and flavin adenine dinucleotide (FADH2) as cofactors. PhrA repairs only cyclobutane pyrimidine dimer but not pyrimidine (6-4) pyrimidinone photoproducts. On the basis of these results, the PhrA protein is classified as a class I, HDF- type, CPD DNA photolyase.
KW - Cyclobutane pyrimidine dimer
KW - DNA photolyase
KW - Over-expression plasmid
KW - Synechocystis sp. strain PCC 6803
KW - pSL1211
KW - phrA
UR - https://www.scopus.com/pages/publications/0034094221
U2 - 10.1007/s002030000164
DO - 10.1007/s002030000164
M3 - Article
C2 - 10896222
AN - SCOPUS:0034094221
SN - 0302-8933
VL - 173
SP - 412
EP - 417
JO - Archives of Microbiology
JF - Archives of Microbiology
IS - 5-6
ER -