Open AccessActivation and transfer of novel synthetic 9‐substituted sialic acids
Author(s) -
GROSS Hans Jürgen,
BÜNSCH Almuth,
PAULSON James C.,
BROSSMER Reinhard
Publication year - 1987
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1987.tb13458.x
Subject(s) - yield (engineering) , chemistry , high performance liquid chromatography , sialic acid , enzyme , glycoside , stereochemistry , substrate (aquarium) , affinity chromatography , transferase , sepharose , chromatography , thiobarbituric acid , acetylglucosamine , atp synthase , biochemistry , biology , ecology , materials science , lipid peroxidation , metallurgy
In this report several NeuAc analogues differently modified at position C‐9 were tested as substrates for CMPsialic acid synthase from bovine brain: the hydroxy group at C‐9 was replaced by an amino, acetamido, benzamido, hexanoylamido and azido group. The synthase was partially purified by chromatography on CDP‐hexanolamine –Sepharose. CMP‐glycosides synthesized were measured by analytical HPLC at 275 nm. Each NeuAc analogue was activated to the respective CMP‐glycoside: K m ‐values varied from 0.8 mM to 4.6 mM, the K m for NeuAc was 1.4 mM. Thus affinity of the enzyme was influenced only moderately by chemical modification at C‐9. CMP‐glycosides were synthesized on a preparative scale with good yield and characterized by analytical HPLC. In addition, 500‐MHz 1 H‐NMR data of CMP‐9‐amino‐NeuAc and CMP‐9‐acetamido‐NeuAc were obtained. Each CMP‐activated NeuAc analogue was a suitable donor substrate for Galβ1‐4GlcNAc α2,6‐sialyl‐transferase from rat liver. Transfer was determined by the thiobarbituric acid method and by analytical HPLC at 200 nm. The results demonstrate that synthetic, not naturally occurring, non‐labelled NeuAc analogues can be incorporated into glycoprotein with high yield.