Reconstitution of an Escherichia coli repair endonuclease activity from the separated uvrA+ and uvrB+/uvrC+ gene products.

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Abstract
An in vitro complementation assay was used for partial purification of the uvrA+, uvrB+ and uvrC+ gene products from E. coli. The uvrB+ and uvrC+ products cochromatograph on DEAE-cellulose and are completely resolved from the uvrA+ product, which was further purified by phosphocellulose chromatography of the nonadsorbed protein fraction from the DEAE-cellulose. Neither the uvrB+/uvrC+ nor the uvrA+ product shows appreciable endonuclease activity on UV-irradiated DNA when tested separately. These factors complement each other to yield an ATP-dependent endonuclease activity specific for UV-irradiated DNA. Gel filtration experiments with the partially purified proteins indicate that the functional uvrA+ gene product has a MW of 100,000. The uvrB+ gene product has an apparent MW of 70,000, but it is presently unclear if this is the size of the uvrB+ product alone or the size of a complex of the uvrB+ and uvrC+ gene products.