Mechanistic studies on shikimate pathway enzymes

酶能使化学反应加速一百多万倍。我们感兴趣的是了解酶内的氨基酸残基是如何带来这种速率加速的。一些残基参与结合，另一些参与化学反应，而其他基团则参与两者。我们的主要重点是参与酪氨酸生物合成的酶。所研究的酶，一个突变酶和一个脱氢酶，在这个途径中催化两个连续的反应。在一些生物体中，这些反应是在单独的多肽上催化的，而在包括大肠杆菌在内的其他细菌中，这种酶属于一种叫做双功能蛋白的特殊类别；在一个多肽上催化两个反应。我们感兴趣的是了解这些酶催化的反应是如何单独工作的，以及描述这两种活动之间的关系。为了获得关于其中一个反应的进一步信息，我们最近从一个超嗜热细菌中解出了单功能脱氢酶的结构。该结构可用于指导诱变研究，探索该酶的作用机制和酪氨酸调控模式。将结果与双功能酶的类似研究进行比较，可以帮助我们深入了解这两个位点的相互依赖性。这些酶催化的反应是植物和微生物所特有的。了解特定的酶残基如何与抑制剂结合，可以设计出更好的除草剂和抗菌剂。

Enzymes can accelerate chemical reactions by more than a million-fold.  We are interested in understanding how amino acid residues within enzymes bring about this rate acceleration.  Some residues are involved in binding, others participate in the chemistry of the reaction, while other groups are involved in both. Our main focus is on enzymes involved in the biosynthesis of tyrosine. The enzymes under study, a mutase and a dehydrogenase, catalyze two consecutive reactions in this  pathway.  In some organisms, the reactions are catalyzed on separate polypeptides while in other bacteria, including E. coli the enzyme belongs to a special class called bifunctional proteins; two reactions are catalyzed on a single polypeptide.  We are interested in understanding how each of these enzyme-catalyzed reactions work separately, as well as delineating the relationship between the two activities.   In order to gain further information about one of the reactions in particular, we have recently solved the structure of a monofunctional dehydrogenase from a hyperthermophilic bacterium.  This structure is used to guide mutagenesis studies to probe the enzyme's mechanism of action and mode of regulation by tyrosine. Comparison of results with analogous studies on the bifunctional enzyme, for which there is no structure, help us gain insight into the interdependence of the two sites. The reactions catalyzed by these enzymes are specific to plants and microoganisms.  Knowledge of how specific enzymic residues bind to inhibitors could lead to the design of improved herbicides and antimicrobial agents.