- A FRET probe for cell-based imaging of ganglioside-processing enzyme activity and high-throughput screening
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Gangliosides are important signaling molecules in the cell membrane and are processed by several enzymes. Deficiencies in these enzymes can cause human lysosomal storage diseases. Building an understanding of the pathways of glycosphingolipid catabolism requires methods for the analysis of these enzymatic activities A GM3-derived FRET probe was synthesized chemoenzymatically for the detection and quantitation of a range of ganglioside-degrading enzymes, both in cell lysates and in living cells. This is the first substrate that enables the ratiometric fluorogenic assay of sphingolipid ceramide N-deacylase and endoglycoceramidase and can detect and localize neuraminidase activity in living cells. It is therefore a valuable tool for building a better understanding of membrane-confined enzymology. It also enables the robust and reliable assay of ganglioside-degrading enzymes in a microtiter plate, thus opening the door to screening for novel or engineered biocatalysts or for new inhibitors.
- Yang, Guang-Yu,Li, Caishun,Fischer, Michael,Cairo, Christopher W.,Feng, Yan,Withers, Stephen G.
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- Glycosphingolipid synthesis employing a combination of recombinant glycosyltransferases and an endoglycoceramidase glycosynthase
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Glycosynthase mutants of Rhodococcus sp. endo-glycoceramidase II efficiently synthesize complex glycosphingolipids. Glycosyl fluoride donors may be assembled via sequential glycosyltransferase-catalysed glycosylation of lactosyl fluoride. Alternatively, lactosyl fluoride may be coupled to sphingosine prior to subsequent glycosylation steps.
- Rich, Jamie R.,Cunningham, Anna-Maria,Gilbert, Michel,Withers, Stephen G.
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- Glycosynthase-mediated synthesis of glycosphingolipids
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Glycosphingolipids play crucial roles in virtually every stage of the cell cycle, and their clinical administration has been proposed as a treatment for Alzheimer's, Parkinson's, stroke, and a range of other conditions. However, lack of supply has severely hindered testing of this potential. A novel glycosynthase-based synthetic strategy is demonstrated, involving a mutant of an endoglycoceramidase in which the catalytic nucleophile has been ablated. This mutant efficiently couples a range of glycosyl fluoride donors with a range of sphingosine-based acceptors in yields around 95%. This technology opens the door to large-scale production of glycosphingolipids and, thus, to clinical testing. Copyright
- Vaughan, Mark D.,Johnson, Karl,DeFrees, Shawn,Tang, Xiaoping,Warren, R. Antony J.,Withers, Stephen G.
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- A Diversity-Oriented Strategy for Chemoenzymatic Synthesis of Glycosphingolipids and Related Derivatives
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A diversity-oriented strategy combining enzymatic glycan assembly and on-site lipid remodeling via chemoselective cross-metathesis and N-acylation was developed for glycosphingolipid (GSL) synthesis starting from a common, simple glycoside. The strategy was verified with a series of natural GSLs and GSL derivatives and showed several advantages. Most notably, it enabled two-way diversification of the glycan and lipid, including introduction of designed molecular tags, to provide functionalized GSLs useful for biological studies and applications.
- Li, Qingjiang,Jaiswal, Mohit,Rohokale, Rajendra S.,Guo, Zhongwu
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- One-Pot Enzymatic Synthesis and Biological Evaluation of Ganglioside GM3 Derivatives as Potential Cancer Immunotherapeutics
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Cancer is a leading cause of death worldwide. Recent research studies have revealed that GM3 derivatives have considerable promise as potential therapeutic agents for cancer. To discover novel GM3 derivatives as potential antitumor agents, a one-pot enzymatic synthesis was established, yielding 14 GM3 derivatives in high total yields (22-41%). Subsequently, the inhibitory activities of GM3 derivatives were assessed by wound-healing assays and Transwell assays and tumor-bearing animal models. Among all the GM3 derivatives, N-12 showed excellent migration and invasion inhibitory effects in cells and marked antitumor activity in C57BL/6 mice. The subsequent analysis of cancer tissues and serum samples revealed that N-12 induces tumor inhibition, which was closely related to immune response. Taken together, N-12 can be further developed as an effective therapeutic for the treatment of cancer. An RNA-sequencing (RNA-seq) analysis was then performed and indicated that the antitumor mechanism of N-12 involved focal adhesion and ECM-receptor interaction signaling pathways.
- Wang, Juntao,Lu, Dan,Sun, Ran,Lei, Shuwen,Luo, Shuhua,Dang, Xin,Zhang, Yang,Yuan, Chang,Zhang, Yong,Wu, Jinhong,Yang, Guangyu,Fu, Lei,Jiang, Faqin
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p. 1883 - 1897
(2022/02/10)
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- Chemoenzymatic synthesis and biological evaluation of ganglioside GM3 and lyso-GM3 as potential agents for cancer therapy
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A highly efficient chemoenzymatic method for synthesizing ganglioside GM3 and lyso-GM3 was reported here. Enzymatic extension of the chemically synthesized lactosyl sphingosine using efficient one-pot multienzyme (OPME) reaction allowed glycosylation to b
- Li, Tingshen,Wang, Xiaodan,Dong, Peijie,Yu, Peng,Zhang, Yongmin,Meng, Xin
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- Chemoenzymatic Total Synthesis of GM3 Gangliosides Containing Different Sialic Acid Forms and Various Fatty Acyl Chains
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Gangliosides are sialic acid-containing glycosphingolipids that have been found in the cell membranes of all vertebrates. Their important biological functions are contributed by both the glycan and the ceramide lipid components. GM3 is a major ganglioside
- Yu, Hai,Gadi, Madhusudhan Reddy,Bai, Yuanyuan,Zhang, Libo,Li, Lei,Yin, Jun,Wang, Peng G.,Chen, Xi
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p. 8672 - 8682
(2021/07/20)
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- Ganglioside GM3 and/or analogue thereof, synthesis method and application of ganglioside GM3 and/or analogue thereof
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The invention relates to a method for synthesizing ganglioside GM3 and/or an analogue thereof. The method includes the following steps: (1) selecting a lactose donor represented by a general formula Iand/or an analogue thereof and selecting a sphingosine
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- GLYCOSYLATED SPHINGOID BASES AND PRODUCTION THEREOF
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The present invention relates to several novel 1-O-glycosylated sphingoid bases and to a production method thereof, as well as to uses of the 1-O-glycosylated sphingoid bases. Sphingoid bases carrying a vinylogous amide-type protecting group are used here
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Page/Page column 43-44
(2020/01/08)
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- Streamlined chemoenzymatic total synthesis of prioritized ganglioside cancer antigens
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A highly efficient streamlined chemoenzymatic strategy for total synthesis of four prioritized ganglioside cancer antigens GD2, GD3, fucosyl GM1, and GM3 from commercially available lactose and phytosphingosine is demonstrated. Lactosyl sphingosine (LacβS
- Yu, Hai,Santra, Abhishek,Li, Yanhong,McArthur, John B.,Ghosh, Tamashree,Yang, Xiaoxiao,Wang, Peng G.,Chen, Xi
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supporting information
p. 4076 - 4080
(2018/06/12)
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- Highly efficient chemoenzymatic synthesis and facile purification of α-Gal pentasaccharyl ceramide Galα3nLc4βCer
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A highly efficient chemoenzymatic method for synthesizing glycosphingolipids using α-Gal pentasaccharyl ceramide as an example is reported here. Enzymatic extension of the chemically synthesized lactosyl sphingosine using efficient sequential one-pot mult
- Santra, Abhishek,Li, Yanhong,Yu, Hai,Slack, Teri J.,Wang, Peng George,Chen, Xi
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supporting information
p. 8280 - 8283
(2017/07/26)
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- Analogues of glycosphingolipids and glycerolipids suitable for conjugation to gold- and amino-functionalised surfaces
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A general synthesis of analogues of natural lipids, (i.e. lactosylceramide, globotriasylceramide, and phosphatidylcholine) where one of the alkyl chains carries a terminal thiol- or carboxyl functionality, is described. The lipids were prepared by N- or O-acylation of sphingosine or monoacylglycerol derivatives. These lipids are suitable for anchoring to gold- or amino-functionalised surfaces, thus creating mimics of a cell membrane for use in the study of protein-carbohydrate interaction. (C) 2000 Elsevier Science Ltd.
- Ohlsson, J?rgen,Magnusson, G?ran
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p. 9975 - 9984
(2007/10/03)
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