Functional complementation of xeroderma pigmentosum complementation group E by replication protein A in an in vitro system

被引:61
作者
Kazantsev, A [1 ]
Mu, D [1 ]
Nichols, AF [1 ]
Zhao, XD [1 ]
Linn, S [1 ]
Sancar, A [1 ]
机构
[1] UNIV CALIF BERKELEY, DIV BIOCHEM & MOLEC BIOL, BERKELEY, CA 94720 USA
关键词
DNA repair; damage recognition; excision nuclease;
D O I
10.1073/pnas.93.10.5014
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Xeroderma pigmentosum (XP) is caused by a defect in nucleotide excision repair. Patients in the complementation group E (XP-E) have the mildest form of the disease and the highest level of residual repair activity. About 20% of the cell strains derived from XP-E patients lack a damaged DNA-binding protein (DDB) activity that binds to ultraviolet-induced (6-4) photoproducts with high affinity. We report here that cell-free extracts prepared from XP-E cell strains that either lacked or contained DDB activity were severely defective in excising DNA damage including (6-4) photoproducts. However, this excision activity defect was not restored by addition of purified DDB that. In fact, inhibited removal of (6-4) photoproducts by the human excision nuclease reconstituted from purified proteins. Extensive purification of correcting activity from HeLa cells revealed that the correcting activity is inseparable from the human replication/repair protein A [RPA (also known as human single stranded DNA binding protein, HSSB)]. Indeed, supplementing XP-E extracts with recombinant human RPA purified from Escherichia coli restored excision activity. However, no mutation was found in the genes encoding the three subunits of RPA in an XP-E (DDB-) cell line. It is concluded that RPA functionally complements XP-E extracts in vitro, but it is not genetically altered in XP-E patients.
引用
收藏
页码:5014 / 5018
页数:5
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