本論文主要是研究特殊環狀多?對DNA序列的選擇性。環?的胺基酸序列組成以Gly-(X-Pro-Arg-Lys)n-Cys為主。DNase I足跡實驗分析後，發現六種環十四?對135-mer pBR322 DNA的選擇性結合區域為5’-TATTTT-3’ (113-108)、5’-TTTTCT-3’ (111-106)及5’-TATTT-3’ (86-83)。另外，CHyD-14、CNW-14、CH2Y-14及CRHY-14在序列115-102區段有相當寬廣的結合位，推測可能有2-3個環?分子並排結合在DNA的特定部位，而CHyY-14及CMY-14則無此現象。對於248-mer pBS DNA的主要結合區域則落在5’-XTTX-3’的位置。由此可知，環十四?對於DNA的TXTTTT序列存在專一性結合的能力，可辨識連續三到四個胸腺嘧啶 (T)。 環九?CWKK-9對276-mer pBS DNA序列選擇性結合的區域主要是5’-CTAG-3’ (31-34)、5’-TTTTG-3’ (64-68)及5’-TCCCTTTA-3’ (70-77)。而環十?中的CW-10對相同片段DNA的5’-CTAG-3’ (31-34)有明顯的專一性結合。但是，將胺基酸殘基Trp置換為Tyr後的CY-10，則多出了許多強結合位，相對地減少了專一性的能力。 此研究顯示經過XPRK修飾的環?具有序列選擇性的結合能力，期望在未來可做為一有用的調控基因表現之試劑。 This thesis reports DNA sequence-selective binding studies of cyclic peptides containing repeating XPRK motifs. DNase I footprinting studies showed that six 14-mer cyclic peptides possess DNA sequence selective binding on a 135-mer DNA fragment, with major preference on 5’-TATTTT-3’, 5’-TTTTCT-3’ and 5’-TATTT-3’ sequences. Of interest, cyclic peptides CHyD-14, CNW-4, CH2Y-14, and CRHY-14 all possess selective binding on a broad locus around positions 102-115, suggesting that about 2-3 cyclic peptide molecules preferentially bind to these positions in a “side-by-side” pattern. On the other hand, cyclic peptides CHyY-14 and CMY-14 are devoid of this “side-by-side” binding mode. On a 248-mer DNA fragment, these cyclic peptides showed preferential binding on 5’-XTTX-3’ sequences (where X = purines). On a 276-mer DNA fragment, cyclic peptide CWKK-9 showed preferential binding around sequences 5’-CTAG-3’ at positions 31-34, 5’-TTTTG-3’ at positions 64-68, and 5’-TCCCTTTA-3’ at positions 70-77. Cyclic peptide CW-10 showed preferential binding on sequence 5’-CTAG-3’ (31-34). These studies suggest that the cyclic peptides containing repeating XPRK motifs possess sequence selective binding capabilities and further studies may provide potential agents for the modulation of gene expression.
