NEW PATHWAY OF ISOPRENOID BIOSYNTHESIS

类异戊二烯生物合成的新途径

• 批准号: 6309041
• 负责人: HEINZ G FLOSS
• 金额: $ 2.74万
• 依托单位: UNIVERSITY OF CALIF-LOS ALAMOS NAT LAB
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-01 至 2002-01-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6309041
• 关键词: biological products; biomedical resource; proteins

The SIR provided (2S)-N-(2'-O-benzyl-glycoloyl)bomane-10-,2-sultam 11,2- 13C]acetic acid; 20g Recently, a new pathway to isopentenyl pyrophosphate has been discovered which emanates from carbohydrate metabolism. To probe distribution of this new pathway in nature and to elucidate further intermediates in the pathway, we are presently synthesizing 1-deoxy-D-[2,4-13C2]xylulose. Initially, a small quantity for exploratory work will be prepared from ethyl bromo- [ I - I 3C] acetate and [2-13C]acetone, which has been developed by the UW group. In the course of this work it became evident that this route is not very efficient and thus unsuitable for the production of larger quantities of this precursor. A second route has therefore been designed by the SIR, which makes effective use of the labeled synthons they have developed. If experiments on ubiquinone formation in E. coli with the initially synthesized precursor sample demonstrate the feasibility of this approach, a larger quantity of the compound will then be synthsized by the SIR. This precursor will be fed to a variety of biological systems synthesizing isoprenoid compounds and the products will be analyzed by 13C-NMR for the incorporation of 13C and the appearance of 13C_13C coupling, resulting from the rearrangement of the precursor, in the appropriate positions. This will give an overview of the relative importance of the new pathway compared to the classical mevalonate pathway. Next, we will incubate the double labeled substrate with washed cells and cell-free extracts of E. coli directly in the NMR spectrometer and examine the incubations periodically by 1H-detected 13C-NMR, looking specifically for double-quantum transitions, for the appearance of new species in which the two 13C nuclei are connected to each other. Since virtually no other metabolic pathway can give rise to extensive 13C-13C coupling, this will be a very specific probe for the intermediates in the new pathway of isoprenoid biosynthesis. The time course of the appearance and disappearance of new species will indicate their relative position in the reaction sequence, and labeling of the precursor in additional positions will allow the assignment of structures to these intermediates. These in vivo experiments will require high sensitivity to detect multiquantum transitions from very small amounts of transient compounds. This can be achieved on the 750 MHz NMR spectrometer at the Univ. of Wash.

SIR提供了(2S)- n -（2'- o -苄基-甘油基）氨基- 10,2 -苏坦