Hyperthermophilic enzymes suitable for functioning membranes.

将超嗜热酶应用于人造功能膜的最重要的一点是它们的极端稳定性。人工膜的重构已经实现，有许多生物传感器在实际应用中。然而，这些制剂的稳定性并不令人满意。为此，重点研究了以下两种超嗜热酶：a。超嗜热v型atp酶的稳定性和功能研究，其同源膜atp酶已被报道。b.控制蛋白质高级结构的嗜热性伴侣蛋白的分离与功能研究。由于膜atp酶和伴侣蛋白都含有相同的atp酶结构，因此克隆是基于结构的。使用的超嗜热菌是Desulfurococcus和thermococcus .a.通过计算机同源性搜索和x射线晶体学鉴定了超嗜热菌和中嗜热菌v - atp酶之间的稳定残基交换。稳定残基交换最丰富的是Asp - Glu（螺旋稳定），其次是Ser - Ala和Gly - Ala（增强内部疏水性）。对这些基因进行克隆后，分离出伴侣蛋白和消旋酶等伴侣蛋白。伴侣蛋白与嗜热真细菌的HSP70有很大差异。对超嗜热伴侣蛋白的详细分析揭示了其α - β亚基（j.l l. biol .）。[j]。最后，这些研究为J.E.Walker和P.D.Boyer在atp酶方面获得诺贝尔奖（1997）做出了贡献。Walker是我们关于嗜热性atp酶的x射线晶体学分析报告的合著者。Boyer经常引用Ann关于嗜热atp酶的研究成果。学生物化学Rev.of。他将于1998年3月在济济医学院给我们做一个关于这个问题的讲座。

The most important point in the application of hyperthermophilic enzymes for an artificial functional membrane is their extreme stability. The reconstitution of the artificial membrane has already been achieved, and there are many biosensors in practical uses. However, the stability of these preparations is not satisfactory.For this reason, the research was focus on the following two hyperthermophilic enzymes :a.The stability and functional study on the hyperthermophilic V-type ATPase, for which homological membrane ATPases have already been reported. The hyperthermophile used was Desulfurococcus which was compared with mesophiles.b.The isolation and functional study on the hyperthermphilic chaperones which controls the higher structure of proteins.Since both membrane ATPases and chaperones contain common ATPase structure, cloning was based on the structure. The hyperthermophiles used were Desulfurococcus and Thermococcus.a.The identification of the stabilizing residue exchanges between the hyperthermophilic and mesophilic V-ATPases was based on the computerized homology search and the X-ray crystallography. The most abundant stabilizing residue exchange was Asp to Glu (helixstabilizing), and the next were Ser to Ala, and Gly to Ala (enhancing the internal hydrophobicity).b.The chaperones including chaperonin and racemase was isolated after the cloning of these genes. The chaperones were quite different from HSP70 of the eubacterial thermophiles. The detailed analysis of the hyperthermophilic chaperone revealed its alpha beta subunits (J.Mol.Biol.273 : 635,1997).Finally, these studies contributed to Nobel Prize (1997) won by J.E.Walker and P.D.Boyer on the ATPase. Walker is the coauthor of our report on X-ray crystallographic analysis of thermophilic ATPase. Boyer frequently quoted works on thermophilic ATPases in Ann. Rev.of Biochem., and will give us a lecture on this subject in Jichi Medical School in March 1998.