細菌利用DNA 複製重啟之引子合成體 (DNA replication restart primosome) 來重新啟
動DNA 複製對其存活而言已被確認為必需，此蛋白質-核酸複合體包含八個蛋白質有次序
的互相結合於停滯的複製叉上，分別為PriA、PriB、PriC、SSB、DnaB、DnaC、DnaG 與
DnaT。克雷伯氏肺炎桿菌是台灣最常被分離出的病原菌之一，為了對抗被克雷伯氏肺炎桿
菌感染的威脅，阻斷其DNA 複製叉重啟的能力也許可行。這個計畫在兩年的時間已完成
的工作如下：(1) 此八個蛋白質的基因選殖、大量表達、純化與蛋白質晶體之高速篩選。(2)
PriB 與SSB 的單股DNA 結合模式之分析。(3) PriB 與DnaT 蛋白質晶體之獲得。(4) PriB
結晶結構解出至2.07Å。(5) 類黃酮物質能抑制DnaB 活性。(6) PriB 與引子合成體蛋白質-
蛋白質間結合作用分析。(7) DnaB 與其抑制劑galangin 蛋白質複合晶體之獲得。希望這些
結果能逐步引導以結構為依據的標靶藥物設計來針對克雷伯氏肺炎桿菌的DNA 複製重啟
體組裝，尤其是PriB、SSB 與DnaB。
It has been recognized that the ability of DNA replication restart primosome is essential for
bacterial survival. These primosomal proteins, PriA, PriB, PriC, SSB, DnaB, DnaC, DnaG and
DnaT, sequentially associated together and then restart arrested DNA replication forks.
Klebsiella pneumoniae (KP) is one of the most commonly isolated bacterial pathogens in Taiwan.
To combat the threat of KP infections, blocking the restart of replication fork of KP may be
useful. The accomplished experiments of this project are: (1) Gene cloning, expression, and
purification of these primosomal proteins. (2) The analyses of ssDNA binding modes of PriB and
SSB. (3) PriB and DnaT protein crystals are obtained. (4) Crystal structure of PriB has been
solved to 2.07 Å by synchrotron radiation X-ray source at Beamline 13C1 of NSRRC. (5) The
flavonols are found to inhibit the activity of DnaB. (6) Analyses of interactions of PriB with the
primosomal proteins. (7) Crystal of DnaB in complex with the inhibitor galangin. We hope
these results will lead the structure-based drug design to target KP replication restart primosome,
especially PriB, SSB and DnaB.
