Zamir L O, Jung E, Jensen R A
J Biol Chem. 1983 May 25;258(10):6492-6.
A mutant strain of Neurospora crassa blocked in each of the initial steps of tryptophan, tyrosine, and phenylalanine biosynthesis was previously shown to accumulate and secrete prephenate and L-arogenate (Jensen, R.A., Zamir, L.O., St. Pierre, M., Patel, N., and Pierson, D.L. (1977) J. Bacteriol. 132, 896-903). We now report the co-accumulation of yet another compound which was identified (Zamir, L.O., Tiberio, R., Jung, E., and Jensen, R.A. (1982) J. Biol. Chem. (1983) 258, 6486-6491) as the lactam derivative of L-arogenate. This structure, spiro-arogenate, undergoes a facile acid-catalyzed conversion to L-phenylalanine (as does L-arogenate). Since L-arogenate is conveniently quantitated as 5-dimethylaminonapthalene-1-sulfonyl (dansyl)-phenylalanine following acidification and dansylation, the unknown presence of spiro-arogenate may easily lead to overestimation of L-arogenate present in mixtures. Reliable quantitative assays for both L-arogenate and spiro-arogenate in mixtures were designed utilizing [3H]dansyl-chloride and exploiting the inability of the spiro-arogenate molecule to be dansylated in contrast to L-arogenate. The initial appearance of spiro-arogenate during accumulation lagged behind prephenate and L-arogenate, and spiro-arogenate accumulation leveled off after 5 days while prephenate and L-arogenate accumulations continued. It seems likely that spiro-arogenate is derived directly from L-arogenate. Prephenate, L-arogenate, and spiro-arogenate comprised about 70, 15, and 15% fractions of the total accumulation in a representative accumulation experiment designed to maximize spiro-arogenate yields. Modest variations in co-metabolite ratios were obtained under nutritional conditions where carbon source, growth temperature, duration of incubation time, and amino acid additions were experimental variables.
先前已表明，在色氨酸、酪氨酸和苯丙氨酸生物合成的每个初始步骤中受阻的粗糙脉孢菌突变株会积累并分泌预苯酸和L-阿洛酸（詹森，R.A.，扎米尔，L.O.，圣皮埃尔，M.，帕特尔，N.，和皮尔逊，D.L.（1977年）《细菌学杂志》132卷，896 - 903页）。我们现在报告另一种化合物的共同积累，该化合物被鉴定（扎米尔，L.O.，蒂贝里奥，R.，荣格，E.，和詹森，R.A.（1982年）《生物化学杂志》（1983年）258卷，6486 - 6491页）为L-阿洛酸的内酰胺衍生物。这种结构，即螺阿洛酸，易于在酸催化下转化为L-苯丙氨酸（L-阿洛酸也是如此）。由于在酸化和丹磺酰化后，L-阿洛酸可方便地定量为5-二甲基氨基萘-1-磺酰基（丹磺酰）-苯丙氨酸，螺阿洛酸的未知存在可能容易导致对混合物中L-阿洛酸含量的高估。利用[³H]丹磺酰氯并利用螺阿洛酸分子与L-阿洛酸不同不能被丹磺酰化的特性，设计了混合物中L-阿洛酸和螺阿洛酸的可靠定量测定方法。螺阿洛酸在积累过程中的最初出现滞后于预苯酸和L-阿洛酸，并且螺阿洛酸的积累在5天后趋于平稳，而预苯酸和L-阿洛酸的积累仍在继续。螺阿洛酸似乎直接来源于L-阿洛酸。在一个旨在使螺阿洛酸产量最大化的代表性积累实验中，预苯酸、L-阿洛酸和螺阿洛酸分别占总积累量的约70%、15%和15%。在碳源、生长温度、孵育时间和氨基酸添加量为实验变量的营养条件下，共代谢物比例有适度变化。
