Characterization of mevalonate kinase V377I, a mutant implicated in defective isoprenoid biosynthesis and HIDS/periodic fever syndrome.
Characterization of mevalonate kinase V377I, a mutant implicated in defective isoprenoid biosynthesis and HIDS/periodic fever syndrome.
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甲羟戊酸激酶 V377I 的表征，这是一种与类异戊二烯生物合成缺陷和 HIDS/周期性发热综合征有关的突变体。
DOI：
10.1016/s1388-1981(01)00105-6
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
Miziorko,HM
中科院分区：
文献类型：
--
作者：
Ríos,SE;Cho,YK;Miziorko,HM
与脂质代谢缺陷有关的疾病清单最近随着高免疫球蛋白血症D和周期性发热综合征(HID：MIM 260920)的增加而增加，这些疾病与甲氧丙戊酸激酶活性[1，2]和蛋白质[1]水平降低有关。更具体地说，已经提出了V377I替代来解释这种疾病。我们观察到V377在真核间戊酸激酶中似乎并不是一成不变的。原核生物甲氧丙酮酸激酶的相对分子质量较低，并且有几个在真核蛋白的377个残基之前终止。这些观察结果促使我们直接测试V377对活性和蛋白质稳定性的影响，方法是在重组人甲氧戊酸激酶中设计V377I突变。突变蛋白已被分离出来，并进行了动力学分析。与野生型酶相比，V377I只表现出轻微的差异(特别是Km(Mva)的6倍≥膨胀)，这不能解释在HIDS细胞提取液中检测到的甲戊酸激酶活性降低的原因。此外，分离的野生型和V377I酶的热失活(50℃)显示这些蛋白质之间的稳定性几乎没有差异。我们的结论是，单一的V377I替换不太可能解释在HID中观察到的甲氧丙酮酸激酶的稳定性和催化活性。
The list of diseases linked to defects in lipid metabolism has recently been augmented by the addition of hyperimmunoglobulinemia D and periodic fever syndrome (HIDS: MIM 260920), which are correlated with depressed levels of mevalonate kinase activity [1,2] and protein [1]. More specifically, a V377I substitution has been proposed to account for this disease. We observed that V377 appears to be far from invariant in eukaryotic mevalonate kinases. Prokaryotic mevalonate kinases are lower in molecular weight and several terminate prior to residue 377 of the eukaryotic proteins. These observations prompted our direct test of the impact of V377 on activity and protein stability by engineering a V377I mutation in a recombinant human mevalonate kinase. The mutant protein has been isolated and kinetically characterized. In comparison with wild-type enzyme, V377I exhibits only modest differences (notably ≥6-fold inflation of Km(MVA)) that do not account for the diminished mevalonate kinase activity assayed in HIDS cell extracts. Moreover, thermal inactivation (50°C) of isolated wild-type and V377I enzymes demonstrates little difference in stability between these proteins. We conclude that a single V377I substitution is unlikely to explain the observation of depressed mevalonate kinase stability and catalytic activity in HIDS.