<b>Implications of the zinc‐finger motif found in the DNA‐binding domain of the human XPA protein</b>

Morita, Eugene Hayato; Ohkubo, Tadayasu; Kuraoka, Isao; Shirakawa, Masahiro; Tanaka, Kiyoji; Morikawa, Kosuke

doi:10.1046/j.1365-2443.1996.d01-252.x

Cited by 39 publications

(27 citation statements)

References 21 publications

(22 reference statements)

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“…Although the association of XPA-MBD with zinc has been previously shown by atomic absorption mass spectrometry (Morita et al, 1996), this is the first time cobalt associated XPA-MBD has been reported. More importantly, the EXAFS data provide direct evidence that the metal, in both ZnXPA-MBD and CoXPA-MBD, is coordinated to the sulphur atoms of four cysteine residues.…”

Section: Nj Hess Et Almentioning

confidence: 61%

“…Despite substantial indirect evidence for a C4-type metal-binding motif in XPA-MBD (Kuraoka et al, 1996, Morita et al, 1996, no definite proof exists. To unambiguously verify that XPA-MBD possesses a metal-binding motif and to identify the ligands coordinated to the metal, extended X-ray absorption fine structure (EXAFS) spectra were collected on two XPA-MBD samples containing zinc (ZnXPA-MBD) and cobalt (CoXPA-MBD).…”

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confidence: 99%

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Human nucleotide excision repair protein XPA: Extended X‐ray absorption fine‐structure evidence for a metal‐binding domain

Hess¹,

Buchko

Conradson

et al. 1998

Protein Science

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The ubiquitous, multi-enzyme, nucleotide excision repair (NER) pathway is responsible for correcting a wide range of chemically and structurally distinct DNA lesions in the eukaryotic genome. Human XPA, a 31 kDa, zinc-associated protein, is thought to play a major NER role in the recognition of damaged DNA and the recruitment of other proteins, including RPA, ERCC1, and TFIIH, to repair the damage. Sequence analyses and genetic evidence suggest that zinc is associated with a C4-type motif, C10S-X2-C108-X17-C126-X2-C129, located in the minimal DNA binding region of XPA (M98-F219). The zinc-associated motif is essential for damaged DNA recognition. Extended X-ray absorption fine structure (EXAFS) spectra collected on the zinc associated minimal DNA-binding domain of XPA (ZnXPA-MBD) show directly, for the first time, that the zinc is coordinated to the sulfur atoms of four cysteine residues with an average Zn-S bond length of 2.34 k 0.01 A. XPA-MBD was also expressed in minimal medium supplemented with cobalt nitrate to yield a blue-colored protein that was primarily (>95%) cobalt associated (CoXPA-MBD). EXAFS spectra collected on CoXPA-MBD show that the cobalt is also coordinated to the sulfur atoms of four cysteine residues with an average Co-S bond length of 2.33 k 0.02 A.Keywords: DNA repair protein; EXAFS; metalloproteins; nucleotide excision repair; XPA Nucleotide excision repair (NER) is a major cellular pathway for removing many structurally distinct DNA lesions from both prokaryotic and eukaryotic genomes (Friedberg et al

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Section: Nj Hess Et Almentioning

confidence: 61%

mentioning

confidence: 99%

Human nucleotide excision repair protein XPA: Extended X‐ray absorption fine‐structure evidence for a metal‐binding domain

Hess¹,

Buchko

Conradson

et al. 1998

Protein Science

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“…Within the central, DNA binding domain of the XPA protein (M98-F219) is the zinc-binding subdomain and a loop-rich subdomain (Figure 4) [22][23][24]26]. The zinc subdomain contains a zinc finger motif: Cys 105 -X 2 -Cys 108 -X 17 -Cys 126 -X 2 -Cys 129 [22][23][24]26].…”

Section: Discussionmentioning

confidence: 99%

“…The zinc subdomain contains a zinc finger motif: Cys 105 -X 2 -Cys 108 -X 17 -Cys 126 -X 2 -Cys 129 [22][23][24]26]. Replacement of any one of the cysteines by a serine leads to disruption of the zinc finger structure and NER [20,27].…”

Section: Discussionmentioning

confidence: 99%

Domains in the XPA protein important in its role as a processivity factor

Bartels

Lambert

2007

Biochemical and Biophysical Research Communications

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XPA is a protein essential for nucleotide excision repair (NER) where it is thought to function in damage recognition/verification. We have proposed an additional role, that of a processivity factor, conferring a processive mechanism of action on XPF and XPG, the endonucleases involved in NER. The present study was undertaken to examine the domain(s) in the XPA gene that are important for the ability of the XPA protein to function as a processivity factor. Using site-directed mutagenesis, mutations were created in several of the exons of XPA and mutant XPA proteins produced. The results showed that the DNA binding domain of XPA is critical for its ability to act as a processivity factor. Mutations in both the zinc finger motif and the large basic cleft in this domain eliminated the ability of XPA to confer a processive mechanism of action on the endonucleases involved in NER.Keywords xeroderma pigmentosum; XPA protein; DNA repair; processive mechanism of action Though the XPA protein is a key component in NER, its precise function in this repair pathway is not clear. There is evidence that it plays a role in the initial steps of NER, where it is involved in damage recognition/verification [1][2][3]. We have proposed an additional important function for XPA, that of a processivity factor needed to confer a processive mechanism of action on the endonucleases, XPF and XPG, involved in NER [4,5]. Proteins can locate target sites on DNA by two distinctive mechanisms: (1) a processive mechanism in which a protein first binds to a random site on DNA and then translocates to a specific site by a facilitated-diffusion process in which it slides or hops along the DNA; or (2) a distributive mechanism, in which a protein has no affinity for non-target DNA and locates target sites by a random, threedimensional diffusion process [6][7][8][9]. The mechanism utilized by DNA-targeting proteins is extremely important in determining the ability of the protein to properly interact with its target sites on DNA and, for many of these proteins, a processive mechanism of action is essential [6,8] We have previously shown that during repair of UVC light induced cyclobutane pyrimidine dimers in normal human cells, the endonucleases, XPG and XPF, incise DNA at sites of damage using a processive mechanism of action [4,5]. In contrast, these endonucleases in cells from

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“…ICP-AES-The zinc content of the TLS zinc finger-like domain (Fr3-ZnF) was determined with an Optima3000 ICP-AES instrument (PerkinElmer Life Sciences) at the Advanced Instrumentation Center for Chemical Analysis, Ehime University (22).…”

Section: Methodsmentioning

confidence: 99%

Domain Architectures and Characterization of an RNA-binding Protein, TLS

Iko

Kodama

Kasai

et al. 2004

Journal of Biological Chemistry

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116

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Translocated in liposarcoma (TLS) is an importantprotein component of the heterogeneous nuclear ribonucleoprotein complex involved in the splicing of pre-mRNA and the export of fully processed mRNA to the cytoplasm. We examined the domain organization of human TLS by a combined approach using limited proteolysis, matrix-assisted laser desorption ionization time-of-flight mass spectrometry, circular dichroism, inductively coupled plasma atomic emission spectroscopy, and NMR spectroscopy. We found that the RNA recognition motif (RRM) and zinc finger-like domains exclusively form protease-resistant core structures within the isolated TLS protein fragments, while the remaining regions, including the Arg-Gly-Gly repeats, appear to be completely unstructured. Thus, TLS contains the unstructured N-terminal half followed by the RRM and zinc finger-like domains, which are connected to each other by a flexible linker. We also carried out NMR analyses to obtain more detailed insights into the individual RRM and zinc finger-like domains. The 113 Cd NMR analysis of the zinc finger-like domain verified that zinc is coordinated with four cysteines in the C4 type scheme. We also investigated the interaction of each domain with an oligo-RNA containing the GGUG sequence, which appears to be critical for the TLS function in splicing. The backbone amide NMR chemical shift perturbation analyses indicated that the zinc finger domain binds GGUG-containing RNA with a dissociation constant of about 1.0 ؋ 10 ؊5 M, whereas the RRM domain showed no observable interaction with this RNA. This surprising result implies that the zinc finger domain plays a more predominant role in RNA recognition than the RRM domain.The translocated in liposarcoma (TLS) 1 protein, also termed FUS, was first identified in human myxoid and round cell liposarcomas as an oncogenic fusion protein with a stressinduced DNA-binding transcription factor, CCAAT enhancerbinding homologous protein (CHOP, also known as GADD153 or DDIT3) (1, 2). The resultant fusion protein (TLS-CHOP), consisting of the N-terminal half of TLS and the full-length CHOP, appears to act as a potent transcription factor possibly by combining the TLS transactivation activity and the CHOP DNA binding activity. A different type of fusion protein, TLS-ERG (a member of the erythroblast transformation-specific (ETS) family of transcription factors), was subsequently detected in human acute myeloid leukemia (3).The normal TLS, consisting of 526 amino acids with a calculated molecular mass of 53 kDa, belongs to a family including the closely related proteins Ewing's sarcoma (EWS) (4) and TAF II 68 (TATA-binding protein-associated factor) (5). Thus, they are collectively called the TET (TLS, EWS, TAF II 68) family. EWS and TAF II 68 interact with components of the RNA polymerase II complex (5, 6). Moreover sarcoma-associated RNA-binding fly homologue (SARFH), a Drosophila homologue of TLS, is colocalized with the polymerase on active chromatin (7). The N-terminal domain of TLS is involved in transcription...

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Implications of the zinc‐finger motif found in the DNA‐binding domain of the human XPA protein

Cited by 39 publications

References 21 publications

Human nucleotide excision repair protein XPA: Extended X‐ray absorption fine‐structure evidence for a metal‐binding domain

Human nucleotide excision repair protein XPA: Extended X‐ray absorption fine‐structure evidence for a metal‐binding domain

Domains in the XPA protein important in its role as a processivity factor

Domain Architectures and Characterization of an RNA-binding Protein, TLS

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