Chemical Synthesis Of A Ketone Substrate Analogue For The Detection Of Intracellular Sugar Chains

sialylated and O-GlcNAc protein are common post-translation modifications inside the cell glycosylation, which are playing important biological roles in life activities of cells. Their abnormal expression are colsely related with many diseases. Therefore, the detection of intracellular protein modififcations such as sialylated and O-GlcNAc glycosylated, can be used as a reasearch reference for the study of occurrence and development of disease and pathogenesis. The method of oligosaccharide metabolic engineering is a common and useful method in the present work, which could detect the glycosylated protein by introducing the specific active group of ketone to the target glycoprotein. Detection of ketone groups can be achieved for the detection of sialic acid and O-GlcNAc glycosylation modification of the target molecule. Generally, the detections of sialic acid and O-GlcNAc glycosylation modification require a class of keto-sugar substrate analogs including UDP-2-keto-Gal and 2-keto-Man. But these two kinds of ketone substrate analogues are not commercialized, lead to the detection of the glycoylated modification is severely hindered. At present, the synthetic methods of these substrate are generally chemical enzyme assay. Chemical enzymatic method required the expression of enzymes which generally has drawbacks such as substrate recognition and catalytic specificity, expensive cofactor and harsh partial reation conditionsThus, the present study focused on the establishment of an universal chemical method for the synthesis of ketone substrate analogues. The research results are as follows:(1) the synthesis of glycal used one-pot method, without purification by silic-gel column chromatography and reduce the loss of sample to improve toluene solution; the reaction was conducted under the condition of 100?, 10 minutes; the yield was 95%. The method simplifies the steps of the reaction of the iodine in the traditional method, and improves the production rate. (3) Osmium tetroxide and sodium periodate method is used to replace ozone oxidation of carbon carbon double bond, generating keto sugar, which can be reacted at room temperature, doesn't need to keep a -78 ? condition which is so harsh. (4) In the synthesis of UDP-ketone-sugar, the C-1 was converted to be the ammonia ester in order to increase the production rate of phosphorylation, instead of the enzyme method; and this method guaranteed the generality of the method.