結構生物學期中報告

1. 結構生物學期中報告 系級 生物科技所 學生 欒媄竹 學號 D93360006

2. Nat. Struct. Biol. 5 1998 701–706. Solution structure of the DNA- and RPA-binding domain of the human repair factor XPA Takahisa Ikegami1, Isao Kuraoka2, Masafumi Saijo2,Naohiko Kodo2, Yoshimasa Kyogoku3, KosukeMorikawa4, Kiyoji Tanaka2 and Masahiro Shirakawa1 1Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0101, Japan. 2Institute for Molecular and Cellular Biology, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565, Japan. 3Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565, Japan. 4Biomolecular Engineering Research Institute, 6-2-3 Furuedai, Suita, Osaka 565, Japan.

5. PDB ID: 1XPA Asymmetric Unit Ribbons Cylinders

7. 研究內容 • NER是生物體中DNA損害的主要修補系統。在此報告前，只有RPA結構被分析出來，而XPA與RPA在NER起始的步驟中是共同作用於受損的DNA，因此以XPA蛋白為分析對象進行研究，成功的分析出了XPA蛋白與RPA和DNA結合的subdomain。

8. NER

9. Fig. 1 a, Sequence alignment of the central domains of human XPA and other XPAs. The numbering is shown for human XPA. The asterisks indicate the zinc-coordinated cysteine residues. The residues identical to human XPA are boxed. The secondary structure of human XPA is indicated. Important residues discussed in the text are colored (green, basic; yellow, acidic; grey, hydrophobic).

10. Fig. 1 b, Best-fit backbone superpositions ofthe 30 final structures of the central domainof human XPA (residues 98–210) in stereo.The backbone atoms of residues 102–155,163–165 and 180–209 are superimposed.Loops L1 (residues 148–163) and L2 (residues166–179) are colored green. c, Schematic ribbondrawing of the NMR structure of thecentral domain of human XPA in stereo,drawn with the programs MOLSCRIPT andRASTER3D. a-helices are green and b-strands are red. Secondary structure elementsare indicated.

11. Structural statistics1 for the central domain of XPA

12. Fig. 2 Structures of the zinc-containing subdomain.a, Best-fit ackbone superpositions of the30 final structures of the zinc-ontaining subdomain(residues 102–129). The backbone atomsof these residues are superimposed. The sidechains of the residues in the hydrophobic coreare colored red, and those of the zinc-bindingcysteines are colored green.

13. Fig. 2 Structures of the zinc-containing subdomain.b, Stick representationof the zinc-containing subdomain. Thelocations of residues that contribute to theacidic patch in the vicinity of the b-hairpin areindicated in red. The locations of residues thatcontribute to the hydrophobic patch in thevicinity of the helical turn are indicated ingreen.

14. Fig. 2 Structures of the zinc-containing subdomain.c, The hydrophobic core formedbetween the zinc-containing subdomain andthe C-terminal subdomain. The hydrophobicside chains of the zinc-containing subdomainthat form the core are colored green, those ofthe C-terminal subdomain are colored red. Allthe figures were produced with the programMOLMOL.

15. Fig. 3 Structures of the C-terminal subdomain.a, Best-fit backbone superpositionsof the 30 final structures of the C-terminalsubdomain (residues 138–209). Thebackbone atoms of the residues 138–155,163–165 and 180–209 are superimposed.Loops L1 (residues 148–163) and L2(residues 166–179) are colored light blue.The side chains of the residues in thehydrophobic core are colored red. b, Thestick representation of the C-terminalsubdomain. The well conserved residuesthat contribute to the positive charges inthe basic cleft are colored blue. The acidicresidues in helices a2 and a3 are coloredred. Both figures were produced with theprogram MOLMOL.

16. Fig. 4 a, Distribution of the electrostatic potential (displayed with GRASP) on the solvent-accessible surface of the central domain of XPA (residues98–210). Blue corresponds to positive potential and red to negative potential. The presenceof a positively charged cleft is evident in the C-terminalsubdomain. In the zinc-ontaining subdomain negatively charged patches are dominant. b, Mapping of the XPA residues with chemical shift perturbationor broadening effects in the (15N, 1H) HSQC spectra. The residues of which the amide resonances were perturbed upon complex formationwith the cisplatin-damaged oligonucleotide (chemical shift perturbation defined by more than 0.08 p.p.m. for 15N or 0.02 p.p.m. for 1H, or broadeningdefined by peak intensities decreased to <50 % of their original values) are indicated in magenta, and those of which the amide resonancesshowed specific broadening upon complex formation with RPA70181–422 (the peak intensities decreased to <65 % of their original values) are coloredgreen. The molecular orientation is the same asin (a)

17. FUTURE WORK • 以XPA蛋白為例練習課程所學之分析方法。 • 本研究已分析出XPA蛋白與RPA及DNA的結合位置，但仍未見到三者間實際結合的模擬圖形；另外可能繼續分析其他的NER相關蛋白與之結合的狀況。