ヒトマトリックスメタロプロテイナーゼ７およびサーモライシンと8-アニリノナフタレン-1-スルホン酸、ヘパリンおよびコレステロール硫酸の相互作用に関する生化学的解析

京都大学0048新制・課程博士博士(農学)甲第19017号農博第2095号新制||農||1029(附属図書館)学位論文||H27||N4899(農学部図書室)31968京都大学大学院農学研究科食品生物科学専攻(主査)教授 保川 清, 教授 安達 修二, 教授 入江 一浩学位規則第4条第1項該当Doctor of Agricultural ScienceKyoto UniversityDFA

Effects of salts on the interaction of 8-anilinonaphthalene 1-sulphonate and thermolysin.

Neutral salts activate and stabilize thermolysin. In this study, to explore the mechanism, we analyzed the interaction of 8-anilinonaphthalene 1-sulphonate (ANS) and thermolysin by ANS fluorescence. At pH 7.5, the fluorescence of ANS increased and blue-shifted with increasing concentrations (0-2.0 μM) of thermolysin, indicating that the anilinonaphthalene group of ANS binds with thermolysin through hydrophobic interaction. ANS did not alter thermolysin activity. The dissociation constants (Kd) of the complex between ANS and thermolysin was 33 ± 2 μM at 0 M NaCl at pH 7.5, decreased with increasing NaCl concentrations, and reached 9 ± 3 μM at 4 M NaCl. The Kd values were not varied (31-34 μM) in a pH range of 5.5-8.5. This suggests that at high NaCl concentrations, Na(+) and/or Cl(-) ions bind with thermolysin and affect the binding of ANS with thermolysin. Our results also suggest that the activation and stabilization of thermolysin by NaCl are partially brought about by the binding of Na(+) and/or Cl(-) ions with thermolysin