With the long-term widespread use of antibiotics,antimicrobial resistance (AMR)has become one of the major health threats in the world. To solve this problem, research groups worldwide have been making continuous efforts to develop new antibacterial drugs.
Recently,Yu Biao's research groupfrom the School of Chemistry and Materials Science, Hangzhou Institute for Advanced Study (HIAS), UCAS has completed the total synthesis and stereochemical assignment of the Nucleoside Antibiotic A-94964 with a novel structure, making new progress and laying a foundation for the research and development of antibacterial drugs of a certain type. The paper on this research entitled "Total Synthesis and Stereochemistry Assignment of Nucleoside Antibiotic A-94964" has been published online inAngew.Chem.Int.E(href="https://doi.org/10.1002/anie.202200818).
The first author of the paper is Dr. Shao Xiaofei.He graduated from the Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences as a member of Yu Biao's Research Group in 2020, after which he joined the School of Chemistry and Materials Science, HIAS, UCAS as a postdoctoral researcher. This research project was funded bythe National Natural Science Foundation of China, the Ministry of Science and Technology of the People's Republic of China, the Chinese Academy of Sciences (CAS) and Science and Technology Commission of Shanghai Municipality.
The long-term extensive use of antibiotics has made bacterial resistance an increasingly prominent problem, posing a serious threat to public health. Therefore, developing antibacterial drugs with a new mechanism of action has become an urgent task. A-94964 is a novel-structure nucleoside antibiotic isolated fromStreptomyces sp., according to a report byDaiichi Sankyo's scientists in 2008. As the activity study shows, A-94964 can effectively inhibit the activity of the phospho-N-acetylmuramoyl-pentapeptide-transferase (mraY)with IC50values of 1.1 μg/mL. Since MraY, an enzyme catalyzing the synthesis of the peptidoglycan precursor Lipid I in bacteria, is highly conserved among different bacterial species, it is considered a potential target for the development of new antibiotics.
Compared with other natural nucleoside antibiotics, A-94964 contains a unique octose aldehyde acid uridine backbone, which is modified at both the C6' and C7' positions. It is linked to a pyranose hexose at the C6' position and to a hexosamine pyranose via a phosphodiester bond at the C7' position. In addition, there is a glycine unit attached to the amino sugar and extended unsaturated fatty acid chains. However, the previous literature only provides the planar structure of A-94964, without any stereochemical information. In fact, there is 1 chiral center and theoretically 216possible stereoisomers in the backbone and sugar units of A-94964.
Kuzuyama's research group from the University of Tokyo, Japan has provided about 3 mg of A-94964 samples through fermentation and purification. Molinaro and Silipo's research group from the University of Naples Federico II, Italy has speculated the structures of a furanose ring and two pyranose rings using the high-field nuclear magnetic resonance technology. The results show that ribose is in the β-configuration, glucosamine in the α-configuration and mannose in the β-configuration. However, the configuration ofmannoseat the anomeric position is identified based on the value of its carbon-hydrogen coupling constant 1JC1"-H1" (165 Hz), so no solid conclusions are available. Additionally, it is impossible to confirm its stereo-configuration based on NMR data because the C6' and C7' sites are in flexible side chains. Due to the heterogeneity of unsaturated fatty acid chains in natural nucleoside antibiotics, there is also no way to research their chiral centers.
Based on the above-mentioned collaborative research, Yu Biao's Lab has started studying the total synthesis and structural confirmation of A-94964. Their first goal is to conduct the total synthesis offour stereoisomers that contain β-configuration mannose units but have different configurations at C6' and C7' positions.A modular synthesis path has been designed to break down the target compounds into four modules, which are then spliced through stereoselective glycosylation, the H-phosphite method, and selective acylation. First, starting from uridine, the research group has rapidly prepared four key 8-carbon uridine stereoisomeric derivatives through stereoselective alkenylation and Sharpless asymmetric dihydroxylation. Subsequently, the monovalent-gold-catalyzed β-selective glycosylation developed by the research group is used to install mannose at the C6' position. Next, the phosphodiester bond is constructed through the H-phosphite method to obtain fully protected trisaccharides, and then all benzyl protecting groups are removed via transfer hydrogenation. Nucleophilic differences are also leveraged to achieve the selective acylation of amino groups and their linkage with unsaturated fatty acid chains. Finally, four possible stereoisomers of A-94964 are obtained by removing the protecting groups under carefully controlled acidic conditions.
Unfortunately, the NMR data of these four stereoisomers are significantly different from those of natural products. The difference is mainly ascribed to the mannose and octose aldehyde acid units. Through the careful analysis of the carbon spectrum data of the mannose unit, the signals at the C3" and C5" positions are in a low biased field compared with the natural products, while the 1JC1"-H1" value is 156 Hz, much less than that of the natural products at 165 Hz. Therefore, the research group speculates that mannose at the anomeric position may have an α-configuration, rather than the originally designated β-configuration.
Thus, a fully benzyl-protected trichloroacetimidate is used as the donor by replacing the glycosidation method of mannose and the mannose unit is introduced α-selectively using steric-hindrance and the anomeric effect. Subsequently, four stereoisomers containing the α-configuration mannose units are efficiently synthesized upon similar transformation. The correct stereochemistry of natural products is determined through the careful analysis and comparison of NMR data. Eventually, A-94964 is identified as a nucleoside compound containing the C6' (S), C7' (S)-octose aldehyde acid uridine backbone, which is modified with α-mannose at the C6' position and linked to the α-glucosamine via a phosphodiester bond at the C7' position.
Source| School of Chemistry and Materials Science
Typesetter | Tong Sihui
Executive Editor | Jiang Xuchen
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