Tammi M, Ballou L, Taylor A, Ballou C E
J Biol Chem. 1987 Mar 25;262(9):4395-401.
Yeast external invertase is a glycoprotein that exists as a dimer that can associate to form tetramers, hexamers, and octamers (Chu, F., Watorek, W., and Maley, F. (1983) Arch. Biochem. Biophys. 223, 543-555; Esmon, P. C., Esmon, B. E., Schauer, I. E., Taylor, A., and Schekman, R. (1987) J. Biol. Chem., 262, 4395-4401), a process that is facilitated by the attached oligosaccharide chains. We have studied this association by high performance liquid chromatography on a gel filtration matrix, by which procedure wild-type bakers' yeast invertase gives two peaks, and invertase from a core mutant (mnn1 mnn9) of Saccharomyces cerevisiae X2180 gives three peaks. Concentration of an invertase solution by freezing drives the dimers into higher aggregates that, at 30 degrees C, re-equilibrate to a mixture of smaller forms, the composition of which depends on pH, concentration, and time. The invertase from a mutant, mnn1 mnn9 dpg1, which underglycosylates its glycoproteins and produces invertase with 4-7 oligosaccharide chains, forms oligomers of much lower stability than the mnn1 mnn9 invertase, which has 8-11 carbohydrate chains. Both of these mutants release external invertase from the periplasm into the medium during growth, but we conclude that defects in the cell wall structure may be more important in this release than an altered tendency of the invertases to aggregate. Investigation of aggregate formation by electron microscopy revealed that all invertases, including the internal nonglycosylated enzyme, form octamers under appropriate conditions.
酵母外切转化酶是一种糖蛋白，以二聚体形式存在，可缔合形成四聚体、六聚体和八聚体（Chu, F., Watorek, W., and Maley, F. (1983) 《生物化学与生物物理学档案》223, 543 - 555；Esmon, P. C., Esmon, B. E., Schauer, I. E., Taylor, A., and Schekman, R. (1987) 《生物化学杂志》, 262, 4395 - 4401），这一过程由附着的寡糖链促进。我们通过在凝胶过滤基质上进行高效液相色谱研究了这种缔合，通过该方法，野生型面包酵母转化酶产生两个峰，而来自酿酒酵母X2180的核心突变体（mnn1 mnn9）的转化酶产生三个峰。通过冷冻浓缩转化酶溶液会使二聚体形成更高聚集体，在30℃时，这些聚集体会重新平衡为较小形式的混合物，其组成取决于pH、浓度和时间。来自突变体mnn1 mnn9 dpg1的转化酶，其糖蛋白糖基化不足，产生具有4 - 7条寡糖链的转化酶，形成的寡聚体稳定性远低于具有8 - 11条碳水化合物链的mnn1 mnn9转化酶。这两种突变体在生长过程中都会将外切转化酶从周质释放到培养基中，但我们得出结论，细胞壁结构缺陷在这种释放中可能比转化酶缔合倾向的改变更为重要。通过电子显微镜对聚集体形成的研究表明，所有转化酶，包括内部非糖基化酶，在适当条件下都会形成八聚体。
