STUDIES ON FOLIC ACID

我们继续我们对二氢叶酸还原酶(DHFR)的研究，DHFR是 叶酸代谢与一系列重要药物的靶点 它们是维生素叶酸的结构类似物。虽然肝脏 是这种还原酶活性的主要部位，最初的尝试是 研究人肝脏中的DHFR是因为观察到酶 这个组织中的活性极低。然而，人类还原酶 (HrDHFR)现已被詹姆斯博士克隆并在大肠杆菌中表达 俄亥俄州医学院Freisheim分校，并在较大范围内提供 金额。在一次持续的合作中，我正在检查某些激活 人体内一种特殊的巯基的现象和性质 像我之前在DHFR中发现和研究的酶，源于 鸡肉、牛肉、猪肉和绵羊。 到目前为止，在研究中，hrDHFR以一种方式响应-SH修饰 与牛肉还原酶观察到的结果相似。大约2倍的激活 在用对氯汞苯甲酸盐(PCMB)治疗后注意到。其他 有机汞化合物既不表现出活化也不表现出抑制作用。这是为了 与鸡肝还原酶(Cldhfr)相比，后者是由 到目前为止检查过的所有有机汞。此外，四硫酸根和 碘激活clDHFR高达5倍，对hrDHFR无影响。 同样，4-5M尿素的最大瞬时激活只有大约2倍， 而相同浓度的尿素的产率约为ClDHFR的5倍。 这是否是人类所能诱导的最大激活 与观察到的酶活性的急剧增加相比 ClDHFR是当前研究的基础。关联的可能性 这些对不同DHFR结构差异的观察， 特别是在从活性中心移除的-SH区域中 正在探索中。此外，与这些观察结果相关的一个假设 潜在的生理功能正在考虑之中。

We continue our studies on dihydrofolate reductase (DHFR), a key enzyme in folic acid metabolism and the target site for a series of important drugs that are structural analogs of the vitamin, folic acid. Although the liver is the major site for this reductase activity, initial attempts to investigate DHFR in human liver was deterred by the observation that enzyme activity was extremely low in this tissue. However, the human reductase (hrDHFR) has now been cloned and expressed in E. coli by Dr. James Freisheim, Medical College of Ohio and is available in somewhat large amounts. In a continuing collaboration, I am examining certain activation phenomenon and the properties of a specific sulfhydryl group in the human enzyme such as I previously discovered and studied in DHFR derived from the chick, beef, pork, and sheep. In studies, thus far, the hrDHFR responds to -SH modification in a manner similar to that observed with the beef reductase. About 2-fold activation is noted after treatment with p-chloromercuribenzoate (pcmb). Other organic mercurials exhibit neither activation nor inhibition. This is to be compared with the chicken liver reductase (clDHFR) that is activated by all organic mercurials examined to date. In addition, tetrathionate and iodine that activate clDHFR as much as 5-fold have no effect on the hrDHFR. Again, the maximum transient activation by 4-5 M urea is only about 2-fold, whereas the same urea concentration yields about 5-fold with the clDHFR. Whether this is the maximum activation that can be induced in the human enzyme as compared with the dramatic increases in activity observed with the clDHFR is the basis for current studies. The possibilities of relating these observations to differences in the structure of the various DHFRs, particularly in the region of the-SH that is removed from the active site are being explored. In addition, a hypothesis relating these observations to a potential physiological function is under consideration.