- 作者: 鄭建中 ; 簡源宏 ; 羅晴容 ; 程家維
- 作者服務機構: 清華大學生物醫學研究所
- 中文摘要: High mobility group protein (HMG) is known to be involved in the formation of high order structure of chromatin. HMGs with minor-groove binding ability in the AT-rich DNA region play a vital role in controlling gene transcription activity. In this report, a 18-residue HMG-I/DAT1 chimeric peptide, PRGRPKGKTLREPRGRPY, was designed and synthesized containing two repetitive PRGRP units and a linking peptide, KGKTLRE, as a targeted DNA-binding peptide. The segment PRGRP is derived from HMG-I while KGKTLRE is from the DATI peptide. Using gel-mobility shift assay and /sup 32/P-end labeled 27 bp AT-rich DNA, the dissociation constant of this chimeric peptide was found to be 4.7 * 10/sup -6/M, that is, 10/ sup 4/ times stronger than that of the PRGRP segment stand alone (> 10/sup -2/M). In addition, the binding constant was found to increase with the length of AT-rich DNA. The possible DNA binding site of the HMG-I/DAT1 chimeric peptide is determined by foot-printing experiments using a minor-groove cleaving agent ruthenium (III)-Schiff base complex and a 135-bp /sup 32/P-5'-end-labeled DNA restriction fragment of Hind III/Rsa I from plasmid pBR322 DNA. The major pattern protected by the HMG-I/DAT1 chimeric peptide exhibits a preference for 5'-AAAT-3' of the AT-rich region. Therefore, this novel design HMG-I/DAT1 chimeric peptide possesses not only a high affinity to AT-rich DNA but also the sequence-specific binding in the minor groove of DNA, which may further lead to the design of short synthetic peptides for therapeutic applications.
- 英文摘要: --
- 中文關鍵字: Minor Groove Binding; Hmg; Footprinting; Ruthenium; Peptide
- 英文關鍵字: 窄溝結合;停經促性腺激素;腳印分析;釕;胜