Open AccessPurification of the human O 6 ‐methylguanine‐DNA methyltransferase and uracil‐DNA glycosylase, the latter to apparent homogeneity
Author(s) -
MYRNES Bjørnar,
WITTWER Christian Urs
Publication year - 1988
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1988.tb14010.x
Subject(s) - uracil dna glycosylase , dna glycosylase , uracil , dna , biochemistry , chemistry , dna methyltransferase , gel electrophoresis , molecular mass , methyltransferase , polyacrylamide gel electrophoresis , enzyme , microbiology and biotechnology , dna repair , biology , methylation
Uracil‐DNA glycosylase, the enzyme that catalyzes the release of free uracil from single‐stranded and double‐stranded DNA, has been purified 26 600‐fold from HeLa S 3 cell extracts. The enzyme preparation was essentially homogeneous as judged by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The native enzyme is a small monomeric protein of molecular mass 29 kDa. A minor uracil‐DNA glycosylase preparation was also obtained in the final chromatographic step. This preparation is homogeneous with a molecular mass of 29 kDa and may represent the mitochondrial enzyme. This report also presents a 700‐fold purification of HeLa S 3 cell O 6 ‐methylguanine‐DNA methyltransferase. The glycosylase and methyltransferase showed very similar chromatographic properties. The report indicates that the lability of the methyltransferase upon purification may be a consequence of the total separation of the two DNA repair enzymes or of the possibility that some other stabilizing factor is involved.