汞和铅对人胎盘碱性磷酸酶活力与构象的影响

The effect of mercury and lead on catalytic activity and conformation of human placental alkaline phosphatase

目的： 研究汞和铅对人胎盘耐热性碱性磷酸酶（ ＨＰＡＰ） 的作用，以探讨汞和铅影响胎儿发育的分子机制。方法：应用分光光度法分别测定不同浓度氯化汞（ ＨｇＣｌ２） 和硝酸铅［Ｐｂ（ＮＯ３）２］ 存在下的ＨＰＡＰ 活力，用荧光法检测其荧光光谱的变化；用双倒数作图法确定ＨｇＣｌ２ 和Ｐｂ（ ＮＯ３）２ 的抑制类型。结果：ＨｇＣｌ２ 和Ｐｂ（ ＮＯ３）２ 均抑制了ＨＰＡＰ 的活力，并使其荧光强度和发射峰位发生了明显改变，但其作用机制不同。ＨｇＣｌ２ 是ＨＰＡＰ 的混合性抑制剂，其抑制常数为３ ．６０ｍ ｍｏｌＬ；而Ｐｂ（ＮＯ３）２ 是ＨＰＡＰ 的反竞争性抑制剂，其抑制常数为１ ．４０ ｍ ｍｏｌＬ。结论：ＨｇＣｌ２ 和Ｐｂ（ＮＯ３）２ 抑制ＨＰＡＰ 的活力，并使其构象发生改变。

AIM:To study the effect of mercury chloride(HgCl 2) and lead nitrate [Pb(NO 3) 2] on human placental alkaline phosphatase(HPAP),and the molecular mechanism of effect of Hg and Pb on development of fetus. METHODS: Spectrophotometry was used to measure the HPAP activity and fluorometry was used to observe the change of fluorescence spectra of HPAP under different concentrations of HgCl 2 and Pb(NO 3) 2.The inhibition type of HgCl 2 and Pb(NO 3) 2 on HPAP was determined by using the double reciprocal plot.RESULTS: The HPAP activity was inhibited and its emission maximum wavelength and fluorescence intensity were obviously changed by HgCl 2 and Pb(NO 3) 2·HgCl 2 is a mixed inhibitor of HPAP and its inhibition constant(Ki) is 3.60 mmol/L.Whereas Pb(NO 3) 2 is an non competitive inhibitor of HPAP and its Ki is 1.40 mmol/L. CONCLUSION: HPAP activity is inhibited and its conformation is changed by HgCl 2 and Pb(NO 3) 2, this may be one of the important molecular mechanisms that mercury and lead affect the development of fetus.