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Differential Labeling of Glycoproteins with Alkynyl Fucose Analogs.

ABSTRACT: Fucosylated glycans critically regulate the physiological functions of proteins and cells. Alterations in levels of fucosylated glycans are associated with various diseases. For detection and functional modulation of fucosylated glycans, chemical biology approaches using fucose (Fuc) analogs are useful. However, little is known about how efficiently each unnatural Fuc analog is utilized by enzymes in the biosynthetic pathway of fucosylated glycans. We show here that three clickable Fuc analogs with similar but distinct structures labeled cellular glycans with different efficiency and protein specificity. For instance, 6-alkynyl (Alk)-Fuc modified O-Fuc glycans much more efficiently than 7-Alk-Fuc. The level of GDP-6-Alk-Fuc produced in cells was also higher than that of GDP-7-Alk-Fuc. Comprehensive in vitro fucosyltransferase assays revealed that 7-Alk-Fuc is commonly tolerated by most fucosyltransferases. Surprisingly, both protein O-fucosyltransferases (POFUTs) could transfer all Fuc analogs in vitro, likely because POFUT structures have a larger space around their Fuc binding sites. These findings demonstrate that labeling and detection of fucosylated glycans with Fuc analogs depend on multiple cellular steps, including conversion to GDP form, transport into the ER or Golgi, and utilization by each fucosyltransferase, providing insights into design of novel sugar analogs for specific detection of target glycans or inhibition of their functions.

SUBMITTER: Ma C

PROVIDER: S-EPMC7503990 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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Differential Labeling of Glycoproteins with Alkynyl Fucose Analogs.

Ma Chenyu C Takeuchi Hideyuki H Hao Huilin H Yonekawa Chizuko C Nakajima Kazuki K Nagae Masamichi M Okajima Tetsuya T Haltiwanger Robert S RS Kizuka Yasuhiko Y

International journal of molecular sciences 20200820 17

Fucosylated glycans critically regulate the physiological functions of proteins and cells. Alterations in levels of fucosylated glycans are associated with various diseases. For detection and functional modulation of fucosylated glycans, chemical biology approaches using fucose (Fuc) analogs are useful. However, little is known about how efficiently each unnatural Fuc analog is utilized by enzymes in the biosynthetic pathway of fucosylated glycans. We show here that three clickable Fuc analogs w ...[more]

PMID: 32825463

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Project description:Fucosylation is one of the most prevalent modifications on N- and O-glycans of glycoproteins, and it plays an important role in various cellular processes and diseases. Small molecule inhibitors of fucosylation have shown promise as therapeutic agents for sickle cell disease, arthritis, and cancer. We describe here the design and synthesis of a panel of fluorinated l-fucose analogs bearing fluorine atoms at the C2 and/or C6 positions of l-fucose as metabolic fucosylation inhibitors. Preliminary study of their effects on cell proliferation revealed that the 6,6-difluoro-l-fucose (3) and 6,6,6-trifluoro-l-fucose (6) showed significant inhibitory activity against proliferation of human colon cancer cells and human umbilical vein endothelial cells. In contrast, the previously reported 2-deoxy-2-fluoro-l-fucose (1) had no apparent effects on proliferations of all the cell lines tested. To understand the mechanism of cell proliferation inhibition by the fluorinated l-fucose analogs, we performed chemoenzymatic synthesis of the corresponding GDP-fluorinated l-fucose analogs and tested their inhibitory activities against the mammalian α1,6-fucosyltransferase (FUT8). Interestingly, the corresponding GDP derivatives of 6,6-difluoro-l-fucose (3) and 6,6,6-trifluoro-l-fucose (6), which are the stronger proliferation inhibitors, showed much weaker inhibitory activity against FUT8 than that of the 2-deoxy-2-fluoro-l-fucose (1). These results suggest that FUT8 is not the major target of the 6-fluorinated fucose analogs (3 and 6). Instead, other factors, such as the key enzymes involved in the de novo GDP-fucose biosynthetic pathway and/or other fucosyltransferases involved in the biosynthesis of tumor-associated glyco-epitopes are most likely the targets of the fluorinated l-fucose analogs to achieve cell proliferation inhibition. To our knowledge, this is the first comparative study of various fluorinated l-fucose analogs for suppressing the proliferation of human cancer and primary endothelial cells required for angiogenesis.

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Differential Labeling of Glycoproteins with Alkynyl Fucose Analogs.

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Differential Labeling of Glycoproteins with Alkynyl Fucose Analogs.

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