幽門螺旋桿菌染色體分離蛋白Soj及Spo0J交互作用之研究

Interaction analysis of the chromosome segregation proteins, Soj and Spo0J from Helicobacter pylori

涂瑋儒(Wei-Ju Tu)

指導教授 ： 孫玉珠

國立清華大學/生命科學院/生物資訊與結構生物研究所/碩士(2014年)

https://doi.org/10.6843/NTHU.2014.00415

摘要

細胞分裂過程中DNA的複製與分配是一項很重要的步驟。Par(partition)即是參與此重要功能的系統，其使複製完成的染色體與質體準確地移動到子細胞內確保遺傳物質數量皆相等。Par系統包含Spo0J及Soj兩種蛋白質與類著絲粒的parS DNA序列。於幽門螺旋桿菌的Par系統中為HpSpo0J與HpSoj兩種蛋白質。Soj是個ATPase。Spo0J是個DNA結合蛋白質會與parS DNA序列結合並且提高Soj水解ATP的速率。利用電泳遷移率變動分析(EMSA)得知 parS DNA (15和17bp)序列與HpSpo0J結合。同樣地運用電泳遷移率變動分析得知55bp的parS DNA與HpSpo0J及HpSoj蛋白質的結合。透過鉬藍試驗(molybdenum-blue assay)得知HpSoj水解ATP速率為0.96 ± 0.3 mole Pi/ mole of Soj．hr。此外,HpSpo0J或parS-55bp個別的參與均會提高HpSoj水解ATP的速率。當HpSpo0J與parS-55bp同時加入時，HpSoj的ATP水解速率提高了45倍。由此猜測HpSpo0J HpSoj parS-55bp三者之間或許會有交互作用。HpSpo0J-HpSoj- parS DNA複合物晶體的培養尚未成功。推測可能原因為HpSpo0J的不穩定與parS-55bp DNA序列太長。

關鍵字

幽門螺旋桿菌

並列摘要

DNA replication and partition played an important role in cell division. To ensure the amounts of genetic material in daughter cells are equal, Par(partition) system makes the replicated DNA moves to daughter cells precisely. Par system contains Spo0J, Soj and centromere-like parS DNA sequence. HpSpo0J and HpSoj are the two proteins from H.pylori for Par system. Soj is an ATPase and binding to nonspecific DNA sequence. Spo0J is a DNA binding protein for parS DNA binding specifically and it can stimulate the ATP hydrolysis of Soj. Using electrophoretic mobility shift assay to detect the parS DNA (15 and 17bp) binding to HpSpo0J. Also, a 55bp parS DNA was used to study the DNA binding to HpSpo0J and HpSoj, respectively by EMSA. The ATPase activity of HpSoj was performed by the molybdenum-blue assay and the ATPase activity of 0.96 ± 0.3 mole Pi/ mole of Soj．hr was determined. HpSpo0J or parS-55bp DNA will stimulate the ATPase activity. The additional of HpSpo0J and parS-55bp DNA mixed the ATPase activity is increased 45 folds the ATPase activity of HpSoj. We speculate that HpSpo0J, HpSoj and parS-55bp may work corporately. Unfortunately, the crystallization of the protein-nucleotide complex, the Spo0J-Soj-parS DNA complex was fail. The fail of crystallization might because the instability of HpSpo0J and the long length of parS-55bp.

並列關鍵字

partition ； Soj ； Spo0J

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