Leucine synthesis in chloroplasts: Leucine/isoleucine aminotransferase and valine aminotransferase are different enzymes in spinach chloroplasts
Leucine synthesis in chloroplasts: Leucine/isoleucine aminotransferase and valine aminotransferase are different enzymes in spinach chloroplasts
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DOI：
10.1016/s0176-1617(97)80176-9
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发表时间：
1997-01-01
影响因子：
4.3
通讯作者：
Schultz, G
中科院分区：
文献类型：
--
作者：
Hagelstein, P;Sieve, B;Schultz, G
这项工作表明，亮氨酸的完全合成定位在叶绿体中。分离纯化的菠菜叶绿体，加入2-氧代异戊酸（3 mmol L（-1）），将[2-C-14]乙酸（2 mmol L（-1））掺入亮氨酸中作为唯一标记的氨基酸。从叶绿体基质中分离出亮氨酸/异亮氨酸氨基转移酶和缬氨酸氨基转移酶作为支链氨基酸合成的末端酶（这两种酶都属于EC 2.6.1类），并纯化约110倍。这些酶的底物特异性差异很大。明显的：亮氨酸/异亮氨酸氨基转移酶的2-氧代异己酸和2-氧代-3-甲基戊酸以及缬氨酸氨基转移酶的2-氧代异戊酸的Km值分别为0.12、0.089和1.13 mmol L（-1）。转氨酶反应的最适pH为8.9。这两种酶竞争性抑制较高浓度的亮氨酸，异亮氨酸和缬氨酸。然而，亮氨酸合成的有效调节是通过微摩尔水平的亮氨酸对2-异丙基苹果酸合酶的灵敏反馈控制而发生的（Hagelstein和Schultz，1993）。关于亮氨酸的合成到叶绿体代谢的整合，途径compering相同的引入底物，乙酰辅酶A和2-oxoisovalerate的问题，进行了讨论。
This work demonstrates that the complete synthesis of leucine is localized in chloroplasts. Isolated, purified spinach chloroplasts, supplied with 2-oxoisovalerate (3 mmol L(-1)), incorporated [2-C-14] acetate (2 mmol L(-1)) into leucine as the only amino acid labelled. A leucine/isoleucine aminotransferase and a valine aminotransferase as terminal enzymes of the branched-chain amino acid synthesis (both enzymes belonging to the EC 2.6.1 class) were isolated from chloroplast stroma and purified about 110-fold. The enzymes differed strongly in their substrate specificities. The apparent: K-m-values for 2-oxoisocaproate and 2-oxo-3-methylvalerate of the leucine/isoleucine aminotransferase and for 2-oxoisovalerate of the valine aminotransferase were 0.12; 0.089 and 1.13 mmol L(-1), respectively. The optimum pH of the aminotransferase reaction was 8.9. Both enzymes were competitively inhibited by higher concentrations of leucine, isoleucine and valine. However, the efficient regulation of leucine synthesis takes place by a sensitive feedback control of 2-isopropylmalate synthase by leucine at a micromolar level (Hagelstein and Schultz, 1993). Concerning the integration of leucine synthesis into chloroplast metabolism, the problem of pathways compering for the same introductory substrates, acetyl-CoA and 2-oxoisovalerate, is discussed.