Sialyltransferases activity with

A HeLa cells cultured in medium containing 5 mM sodium butyrate and 50 ixCi/mL N-[acetyl-sH]-D-mannosamine after 24 hr, one set of cells harvested and medium replaced with fresh medium with and without 0.5 pg/mL cycloheximide on the other two sets, which were harvested after another 19 hr cells then analyzed for radioactive GM3. B Essentially same as A except the cells were cultured in unlabeled medium and assayed for sialyltransferase activity. C same as B except the cells were assayed for GM3-... 

Table I. The Effect of Various Compounds Implicated in the 2-Stage Theory of Carcinogenesis on CMP-NeuAc lactosylceramide Sialyltransferase Activity in Human Epithelial Carcinoma with (KB) Cells...

Table III. Comparison of Serum-induced ODC Activity with CMP-NeuAc lactosylceramide Sialyltransferase Activity in Various Cell Lines...

Kinetic Properties of Sialyltransferases. The sialyl-transferase activities with the endogenous glycoprotein and glycolipid acceptors in the standard assays (15) were linear with time for at least 60 min, while those with the exogenously added GMi and DS-fetuin were linear with time only for about 30 min (Figure 1). Activities were directly proportional to the amount of enzyme added up to 0.75 mg protein/assay (Figure 2). 

Pretreatment of enzyme preparations with trypsin diminished all four sialyltransferase activities (16). The sensitivity to... 

The basis for the multiplicity of the sialyltransferase activities remains to be elucidated. We plan to purify these enzyme species to homogeneity, using isoelectric focusing columns of smaller pH ranges in conjunction with affinity chromatography which has been successfully used to purify the soluble sialyl-transferases from bovine colostrum (57). Possibility exists that the heterogeneity of sialyltransferase activities as observed is due to differences in polypeptide sequences, carbohydrate content, or non-covalent interactions with other membrane components, and these possibilities can be clarified only with highly purified enzyme preparations. 

A continuous sucrose gradient of the total homogenate from young rat brain and electron microscopic examination of these fractions found most of the sialyltransferase activities to be localized in smooth microsomal membrane and Golgi complex derivatives and not associated with synaptosomes. 

Our group has reported a novel multifunctional bacterial enzyme, Pasteurella multocida sialyltransferase (PmSTl). PmSTl is an easy to express extremely soluble, and highly active sialyltransferase. Routinely, about 100 mg of PmSTl can be purified from one liter E. coli cell culture. PmSTl has four different activities including a2,3-sialyltransferase (optimal activity at pH 7.5-9.0), a2,6-sialyltransferase (optimal activity at pH 4.5-7.0), a2,3-sialidase (optimal activity at pH 5.0-5.5), and trans-sialidase (optimal activity at pH 5.5-6.5) activities. Among them, a2,3-sialyltransferase activity is the major function of the PmSTl with a specific activity of 60 U/mg protein (24),... 

We are only beginning to understand the regulation of the expression of sialyltransferase activity. The higher level of sialylation of many tumor cells and increased sialyltransferase activities are well known (see chapter 3 of Vol. 30 and section 10.5 below]. Transformation of FR3T3 cells with the c-Ha-ras oncogene resulted in a marked increase of the expression of 3-galactoside a-2,6-sialyltransferase activity and. 

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