Mechanistic studies on shikimate pathway enzymes

酶可以将化学反应加速一百万倍以上。我们有兴趣了解酶内的氨基酸残基是如何导致这种速度加快的。虽然一些残基参与结合，其他基团参与反应的化学，但其他基团两者都参与。我们主要关注参与酪氨酸生物合成的酶。研究中的酶，一种变位酶和一种脱氢酶，在这一途径中催化两个连续的反应。在一些生物中，这些反应是在单独的多肽上催化的，而在其他细菌中，包括大肠杆菌，这种酶属于一种特殊的类别，称为双功能蛋白；两个反应是在一个多肽上催化的。我们有兴趣了解这些酶催化的反应中的每一个是如何单独工作的，以及描述两个活动之间的关系。我们最近解决了一个超高温细菌的单功能脱氢酶的结构，以获得关于其中一个反应的进一步信息。该结构被用于指导突变研究，以探索酶的作用机制和酪氨酸的调节方式。将结果与没有结构的双功能酶的类似研究进行比较，有助于我们深入了解这两个位点的相互依存关系。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能会导致改进除草剂和抗菌剂的设计。

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.