CRYSTALLOGRAPHIC STUDIES OF TERNARY COMPLEXES OF NMT1P

Nmtlp是一种由455个氨基酸组成的酶，催化这种转移。 肉豆蔻酸对真核细胞N-末端甘氨酸的作用 蛋白质。肉豆蔻酰化被用来调节潜在的可逆 必需的蛋白质-蛋白质和蛋白质-膜相互作用 肉豆蔻酰化蛋白质的生物学功能。 N-肉豆蔻酸化是一种基本的细胞过程和蛋白质， 肉豆蔻酰化包括蛋白酪氨酸激酶，磷酸酶， 异三聚体G蛋白、结构蛋白和非结构蛋白 包括艾滋病毒在内的多种病毒的蛋白质。来自病原体的NMT 真菌也是抗真菌药物设计的重要靶点。我们 最近解决了Nmtlp的结构与一个三元络合物 使用收集的MAD数据的肉豆蔻酰辅酶A和多肽底物类似物 在BNL(Bhatnagar等人)(1998)，《自然结构生物学》，印刷中)。 这种三元复杂结构揭示了 确定酶的底物特性并调节有序的 基材和产品的绑定和释放。这种结构还包括 提出了一种新的催化机制，包括去质子化 多肽底物的N-末端氨基被酶 C-末端主链羧酸盐。我们现在有了同样的水晶 酶结合到相同的肉豆蔻酰辅酶A类似物，但与一个天然的 多肽底物(晶体n 1)。此外，另一个三元络合物 与结合的天然多肽抑制物(晶体n 2)。解开含有天然多肽的复合体的结构将 进一步阐明该酶催化作用的新机制 这是我们在初始结构的基础上提出的。晶体n 1和n 2 空间组的衍射角分别为3.5E和3.3E P3121，单位晶胞尺寸a=b=103E，c=108E 如果数据分辨率不高，同步辐射显然会 对这个项目是有利的。

Nmtlp is an enzyme of 455 amino acids that catalyzes the transfer of myristate to the N-terminal glycine of cellular eukaryotic proteins. Myristoylation is used to mediate potentially reversible protein-protein and protein-membrane interactions that are necessary for the biological function of myristoylated proteins. N-myristoylation is an essential cellular process and proteins that become myrisloylated include protein tyrosine kinases, phosphatases, heterotrimeric G-proteins, as well as structural and non-structural proteins of numerous viruses including HIV. Nmts from pathogenic fungi are also important targets for design of antifungal drugs. We have recently solved the structure of Nmtlp in a ternary complex with a myristoylCoA and a peptide substrate analog using MAD data collected at BNL (Bhatnagar et al. (1998), Nature Structural Biology, in press). This ternary complex structure reveals the structural features that define the enzyme's substrate specificities and regulate the ordered binding and release of substrates and products. This structure also suggests a novel catalytic mechanism which involves deprotonation of the N-terminal ammonium of a peptide substrate by the enzyme's C-terminal backbone carboxylate. We now have crystals of the same enzyme bound to the same myristoylCoA analog but with one natural peptide substrate (crystal n 1). In addition, another ternary complex with a bound natural peptide inhibitor has been generated (crystal n 2). Solving the structure of a complex with a natural peptide will further delineate the novel mechanism of catalysis by this enzyme which we proposed based on the initial structure. Crystal n 1 and n 2 diffracted to 3.5 E and 3.3 E, respectively, were both in space group P3121 with unit cell dimensions a = b = 103 E, c = 108 E. Given the poor resolution of the data, synchrotron radiation would clearly be advantageous to this project.