Mammalian DNA nucleotide excision repair reconstituted with purified protein components
Nucleotide excision repair is the principal way by which human cells remove UV damage from DNA. Human cell extracts were fractionated to locate active components, including xeroderma pigmentosum (XP) and ERCC factors. The incision reaction was then reconstituted with the purified proteins RPA, XPA,...
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| Published in | Cell Vol. 80; no. 6; pp. 859 - 868 |
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| Main Authors | , , , , , , , , , |
| Format | Journal Article |
| Language | English |
| Published |
United States
Elsevier Inc
24.03.1995
Elsevier |
| Subjects | |
| Online Access | Get full text |
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| Abstract | Nucleotide excision repair is the principal way by which human cells remove UV damage from DNA. Human cell extracts were fractionated to locate active components, including xeroderma pigmentosum (XP) and ERCC factors. The incision reaction was then reconstituted with the purified proteins RPA, XPA, TFIIH (containing XPB and XPD), XPC, UV-DDB, XPG, partially purified ERCC1/XPF complex, and a factor designated IF7. UV-DDB (related to XPE protein) stimulated repair but was not essential. ERCC1- and XPF-correcting activity copurified with an ERCC1-binding polypeptide of 110 kDa that was absent in XP-F cell extract. Complete repair synthesis was achieved by combining these factors with DNA polymerase ε, RFC, PCNA, and DNA ligase I. The reconstituted core reaction requires about 30 polypeptides. |
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| AbstractList | Nucleotide excision repair is the principal way by which human cells remove UV damage from DNA. Human cell extracts were fractionated to locate active components, including xeroderma pigmentosum (XP) and ERCC factors. The incision reaction was then reconstituted with the purified proteins RPA, XPA, TFIIH (containing XPB and XPD), XPC, UV-DDB, XPG, partially purified ERCC1/XPF complex, and a factor designated IF7. UV-DDB (related to XPE protein) stimulated repair but was not essential. ERCC1- and XPF-correcting activity copurified with an ERCC1-binding polypeptide of 110 kDa that was absent in XP-F cell extract. Complete repair synthesis was achieved by combining these factors with DNA polymerase epsilon, RFC, PCNA, and DNA ligase I. The reconstituted core reaction requires about 30 polypeptides. Nucleotide excision repair is the principal way by which human cells remove UV damage from DNA. Human cell extracts were fractionated to locate active components, including xeroderma pigmentosum (XP) and ERCC factors. The incision reaction was then reconstituted with the purified proteins RPA, XPA, TFIIH (containing XPB and XPD), XPC, UV-DDB, XPG, partially purified ERCC1/XPF complex, and a factor designated IF7. UV-DDB (related to XPE protein) stimulated repair but was not essential. ERCC1- and XPF-correcting activity copurified with an ERCC1-binding polypeptide of 110 kDa that was absent in XP-F cell extract. Complete repair synthesis was achieved by combining these factors with DNA polymerase ε, RFC, PCNA, and DNA ligase I. The reconstituted core reaction requires about 30 polypeptides. |
| Author | Shivji, Mahmud K.K Podust, Vladimir N Protić, Miroslava Aboussekhra, Abdelilah Biggerstaff, Maureen Vilpo, Juhani A Moncollin, Vincent Wood, Richard D Hübscher, Ulrich Egly, Jean-Marc |
| Author_xml | – sequence: 1 givenname: Abdelilah surname: Aboussekhra fullname: Aboussekhra, Abdelilah organization: Imperial Cancer Research Fund Clare Hall Laboratories South Mimms Herts EN6 3LD England – sequence: 2 givenname: Maureen surname: Biggerstaff fullname: Biggerstaff, Maureen organization: Imperial Cancer Research Fund Clare Hall Laboratories South Mimms Herts EN6 3LD England – sequence: 3 givenname: Mahmud K.K surname: Shivji fullname: Shivji, Mahmud K.K organization: Imperial Cancer Research Fund Clare Hall Laboratories South Mimms Herts EN6 3LD England – sequence: 4 givenname: Juhani A surname: Vilpo fullname: Vilpo, Juhani A organization: Department of Clinical Chemistry Tampere University Hospital Tampere, Finland – sequence: 5 givenname: Vincent surname: Moncollin fullname: Moncollin, Vincent organization: lnstitut de Génétique et de Biologie Moléculaire et Cellulaire 67404 Illkirch Cédex Strasbourg, France – sequence: 6 givenname: Vladimir N surname: Podust fullname: Podust, Vladimir N organization: Department of Veterinary Biochemistry University Zürich-Irchel Winterthurerstrasse 190 CH-8057 Zürich, Switzerland – sequence: 7 givenname: Miroslava surname: Protić fullname: Protić, Miroslava organization: Section on DNA Replication, Repair and Mutagenesis National Institute of Child Health and Human Development Bethesda, Maryland 20892, USA – sequence: 8 givenname: Ulrich surname: Hübscher fullname: Hübscher, Ulrich organization: Department of Veterinary Biochemistry University Zürich-Irchel Winterthurerstrasse 190 CH-8057 Zürich, Switzerland – sequence: 9 givenname: Jean-Marc surname: Egly fullname: Egly, Jean-Marc organization: lnstitut de Génétique et de Biologie Moléculaire et Cellulaire 67404 Illkirch Cédex Strasbourg, France – sequence: 10 givenname: Richard D surname: Wood fullname: Wood, Richard D organization: Imperial Cancer Research Fund Clare Hall Laboratories South Mimms Herts EN6 3LD England |
| BackLink | https://www.ncbi.nlm.nih.gov/pubmed/7697716$$D View this record in MEDLINE/PubMed https://hal.science/hal-03831462$$DView record in HAL |
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| SubjectTerms | Animals Biochemistry, Molecular Biology DNA Damage DNA Ligase ATP DNA Ligases - metabolism DNA Polymerase II DNA Repair DNA Replication DNA-Binding Proteins - isolation & purification DNA-Binding Proteins - metabolism DNA-Directed DNA Polymerase - metabolism Endonucleases HeLa Cells Humans Life Sciences Mammals Plasmids Proliferating Cell Nuclear Antigen - metabolism Proteins - isolation & purification Proteins - metabolism Ultraviolet Rays Xeroderma Pigmentosum Group A Protein |
| Title | Mammalian DNA nucleotide excision repair reconstituted with purified protein components |
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