Identification and isolation of damage-recognition protein complexes from zebrafish(Danio rerio) early embryos.
| Autor: | C.Y. Shih, 施錦毅 |
|---|---|
| Rok vydání: | 2002 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 90 The expression of distorted DNA-binding factors was studied in developing zebrafish using UV-damaged DNA as the binding target. A strong and high-shifting binding activity was detected in the extracts of zebrafish embryo (12 h after fertilization), and the expression of the activity dramatically decreased in 60 to 84-h-old zebrafish. Unlike the embryonic extract, the extracts of 84-h-old larvae produced only low-shifting binding complexes when incubated with a CPD-specific probe. The binding of 12-hr-old and 84-hr-old extracts to CPD- and 6-4PPs under various concentration of NaCl supported the possibility that different damage-recognition protein were expressed in zebrafish according to their developmental condition. A competitive binding assay indicated that both CPDs and 6-4PPs were recognized by the same binding activity expressed in 12-h-old zebrafish. The binding activity expressed in embryos was apparently unrelated to human DNA repair damage-recognition proteins XPA、RPA-70 and RPA-32, since those polypeptides recognized by an anti-human XPA、RPA-70 and RPA-32 antibody were detected only in 84-h-old zebrafish extracts. Identification of one 50.2 and one 29-kDa UV-damage-DNA binding factor in 12-hr-old zebrafish extracts by EMSA linked to SDS-PAGE and determined by MALDI TOF analysis, indicated that the 29-kDa and 50.2-kDa polypeptides are high homologous to zebrafish fast skeletal muscle troponin T and vitellogenin. UV-binding factors found in 12-h-old zebrafish embryos may be involved in processing developmental stage-specific DNA structures similar to UV-damage DNA. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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