Structural basis for the recruitment of ERCC1-XPF to nucleotide excision repair complexes by XPA

Abstract

The nucleotide excision repair (NER) pathway corrects DNA damage caused by sunlight, environmental mutagens and certain antitumor agents. This multistep DNA repair reaction operates by the sequential assembly of protein factors at sites of DNA damage. The efficient recognition of DNA damage and its repair are orchestrated by specific protein-protein and protein-DNA interactions within NER complexes. We have investigated an essential proteinprotein interaction of the NER pathway, the binding of the XPA protein to the ERCC1 subunit of the repair endonuclease ERCC1-XPF. The structure of ERCC1 in complex with an XPA peptide shows that only a small region of XPA interacts with ERCC1 to form a stable complex exhibiting submicromolar binding affinity. However, this XPA peptide is a potent inhibitor of NER activity in a cellfree assay, blocking the excision of a cisplatin adduct from DNA. The structure of the peptide inhibitor bound to its target site reveals a binding interface that is amenable to the development of small molecule peptidomimetics that could be used to modulate NER repair activities in vivo.

Multiple DNA Binding Domains Mediate the Function of the ERCC1-XPF Protein in Nucleotide Excision Repair

“…Our studies allow us to propose how multiple interactions between ERCC1-XPF, XPA, and DNA control incision 5Ј to the damage during NER. ERCC1-XPF is recruited to NER complexes by interaction of the central domain of ERCC1 with XPA, in the context of a pre-incision complex consisting of XPA, RPA, TFIIH, and XPG (10,33). Our cellular studies suggest that DNA binding of ERCC1-XPF is not required to localize the endonuclease to NER complexes (Fig.…”

“…Subsequently, TFIIH verifies the presence of the lesion and opens up the DNA helix, allowing the formation of a pre-incision complex containing the endonuclease XPG, the singlestranded DNA binding protein RPA and the architectural protein XPA (7)(8)(9). Finally, the second endonuclease, ERCC1-XPF is recruited to the pre-incision complex (10,11) and incises the DNA 5Ј to the lesion, triggering initiation of repair synthesis, incision 3Ј to the lesion by XPG, completion of repair synthesis and ligation (12)(13)(14).…”

“…The active site with the conserved nuclease motif is located adjacent to the HhH domain in XPF (30). The central domain of ERCC1 is structurally homologous to the XPF nuclease domain (29,31), however, instead of the active site rich in acidic residues, it contains a groove lined with basic and aromatic residues that interact with the XPA protein, connecting ERCC1-XPF to the NER machinery (10,(32)(33)(34). XPF, the larger of the two proteins, contains an N-terminal SF2-helicase like domain (HLD), which has lost the ability to bind ATP and to unwind duplex DNA (35).…”

Characterization of Hrq1-Rad14 Interaction in Saccharomyces cerevisiae

“…Rad1-Rad10 복합체와 Rad2는 각각 손상 부위의 5' 쪽과 3' 쪽을 자른다 (Guzder et al, 2006;Tsodikov et al, 2007;Mardiros et al, 2011). 대장균의 RecQ helicase에서 그 이름을 따온 RecQ helicase family는 모든 생물체에 잘 보존되어 있으며 유전체의 안정성을 유지하기 위하여 다양한 역할을 한다.…”

Papel das proteínas XPD e DNA polimerase eta nas respostas de células humanas a danos no genoma

“…XPG (XPG/ERCC5), além de essencial para a incisão, também tem um papel de grande importância na estabilização do complexo TFIIH (Ito et al, 2007). O heterodímero XPF-ERCC1 é composto pelas subunidades XPF (103 kDa, produto do gene XPF/ERCC4) e ERCC1 (31 kDa), sendo que ambas são necessárias para a estabilização do complexo (Gaillard e Wood, 2001); XPF contém o domínio catalítico de nuclease, enquanto ERCC1 é necessário para a interação com XPA e outras proteínas de NER (Tsodikov et al, 2007). O complexo XPF-ERCC1 participa não só da via de NER mas também de outros processos de reparo, como o reparo de DSBs (Ahmad et al, 2008) e ICLs (Niedernhofer et al, 2004), além de ser essencial para a viabilidade e desenvolvimento nos seres humanos (Ahmad et al, 2010).…”