Stereoselectivity of the Chinese hamster ovary cell sialidase: sialoside hydrolysis with overall retention of configuration

Glycobiology
Y H KaoT G Warner

Abstract

The stereochemical course of enzymatic hydrolysis by the soluble sialidase from Chinese hamster ovary cells, expressed as a recombinant protein in insect Sf9 cells, was determined using proton nuclear magnetic resonance spectroscopy. 4-Methyl umbelliferyl-N-acetyl neuraminic acid was employed as substrate, and the stereoselectivity of the enzyme catalysis was ascertained by monitoring the H3 axial and equatorial protons of the sialic acid product over the reaction course. At both high (3 U) and low concentrations (1 U) of the enzyme, the alpha anomer of the sialic acid was clearly observed as the initial reaction product. The corresponding beta anomer of sialic acid appeared much later in the reaction, arising from mutarotation of the alpha anomer. Similar studies were also carried out using the Salmonella typhimurium LT 2 sialidase, a protein of similar size and substrate specificity. Both enzymes apparently cleave the alpha linked sialoside substrate with retention of configuration. Based on the observations of a wide variety of other glycohydrolytic enzymes that have shown a strong correlation of the stereoselectivity of catalysis with active site topology (Gebler et al., J. Biol. Chem. 267, 12559-12561, 1992), the results o...Continue Reading

Citations

Jan 2, 2001·Bioorganic & Medicinal Chemistry Letters·J C WilsonM von Itzstein
Jun 19, 2010·Journal of the American Chemical Society·Thomas J Morley, Stephen G Withers
May 5, 2009·The Journal of Biological Chemistry·Thomas J MorleyStephen G Withers
Nov 24, 1999·Glycobiology·E MontiG Borsani
Feb 16, 2005·FEBS Letters·Samia Albouz-AboMark von Itzstein
May 8, 2004·Structure·María Fernanda AmayaPedro M Alzari

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