Characterization of Sialyltransferase‐IV Activity and Its Involvement in the c‐Pathway of Brain Ganglioside Metabolism

Abstract: To characterize the sialyltransferase‐IV activity in brain tissues, the activities of GM1b‐, GD1a‐, GT1b‐, and GQ1c‐synthases in adult cichlid fish and rat brains were examined using GA1, GM1, GD1b, or a cod brain ganglioside mixture as the substrate. The GD1a‐synthase activity in the total membrane fraction from cichlid fish brain required divalent cations such as Mg²⁺ or Mn²⁺ and Triton CF‐54 for its full activity. The V_max value was 1,340 pmol/mg of protein/h at an optimal pH of 6.5, whereas the apparent K_m values for CMP‐sialic acid and GM1 were 172 and 78 µM, respectively. Cichlid fish and rat brains also contained GM1b‐, GT1b‐, and GQ1c‐synthase activities. The ratio of GM1b‐, GD1a‐, and GT1b‐synthase activities in fish brain was 1.00:0.89:1.13, respectively, and in rat brain 1.00:0.60:0.63. Incubation of fish brain membranes with a cod brain ganglioside mixture, which contains GT1c, and [³H]CMP‐sialic acid produced radiolabeled GQ1c. It is interesting that the adult rat brain also contains an appreciable level of GQ1c‐synthase activity despite its very low concentrations of c‐series gangliosides. The GD1a‐ or GQ1c‐synthase activity in fish and rat brain was inhibited specifically by coincubation with the glycolipids that serve as the substrates for other sialyltransferase‐IV reactions. Thus, the GD1a‐synthase activity was inhibited by GA1 and GD1b, but not by LacCer, GM3, or GD3. In a similar manner, the synthesis of GQ1c was suppressed by GA1, GM1, and GD1b, but not by LacCer, GM3, or GD3. The GD1a‐synthase activity directed toward endogenous GM1 was inhibited by GA1 or GT1b, whereas the endogenous GT1b‐synthase activity was suppressed by GA1 or GM1. GA1, GM1, and GD1b did not affect the endogenous GM3‐ and GD3‐synthase activities. These results clearly demonstrate that sialyltransferase‐IV in brain tissues catalyzes the reaction for GQ1c synthesis in the c‐pathway as well as the corresponding steps in the asialo‐, a‐, and b‐pathway in ganglioside biosynthesis.