Assay of DNA photolyase activity in spinach leaves in relation to cell compartmentation-evidence for lack of DNA photolyase in chloroplasts.
Spinach cyclobutane pyrimidine dimer (CPD)-specific DNA photolyase was successfully detected in leaf extracts by an assay system for plant photolyase using an improved enzyme-linked immunosorbent assay (ELISA) which was newly introduced by novel horseradish peroxidase (HRP)-linked CPD specific monoclonal antibodies. The assay system includes two main steps: a photorepair reaction of CPD introduced in substrate DNA and measurement of CPD remained after the photorepair by the improved ELISA. When CPD- induced salmon sperm DNA was used as a substrate, high CPD-photolyase activities were observed in the enzyme fraction prepared from whole spinach leaf extracts, but not from chloroplast extracts. This strongly suggests that spinach CPD-specific photolyases are localized in cell compartments other than chloroplasts.
Hada
M
M
Department of Biology, Faculty of Science, Kobe University, Japan. m-hada@iae.kyoto-u.ac.jp
Hino
K
K
Buchholz
G
G
Goss
J
J
Wellmann
E
E
Shin
M
M
D000818:Animals; D000911:Antibodies, Monoclonal; D002451:Cell Compartmentation; D002736:Chloroplasts / Q000201:enzymology; D004260:DNA Repair; D004255:Deoxyribodipyrimidine Photo-Lyase / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D051379:Mice; D018515:Plant Leaves / Q000201:enzymology; D018724:Spinacia oleracea / Q000201:enzymology*
D000911:Antibodies, Monoclonal; D004255:Deoxyribodipyrimidine Photo-Lyase