Boronic acid lectin affinity chromatography (BLAC). 3. Temperature dependence of glycoprotein isolation and enrichment

Marcell Olajos, Ákos Szekrényes, Peter Hajos, Doug T. Gjerde, András Guttman

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

In this paper, the effect of temperature is investigated on the performance of glycoprotein enrichment by boronic acid lectin affinity chromatography (BLAC). Wheat germ agglutinin and m-aminophenyl boronic acid containing stationary phases were evaluated individually and in a mixed mode using an automated liquid handling robot with an integrated 96-well plate temperature controller. Glycoaffinity enrichment of the model proteins of ribonuclease B and trypsin inhibitor was investigated in the presence of the non-glycosylated proteins of myoglobin (neutral) and lysozyme (basic) at a wide temperature range of 5-65 °C. Our results revealed that glycoaffinity micropartitioning at the temperature of 25 °C provided the highest recovery rate for glycoprotein enrichment. We have also found that a large amount of lysozyme was present in the elution fractions of the m-aminophenyl boronic acid containing micropartitioning columns due to ion-exchange mechanism occurring between the positively charged protein and the negatively charged stationary phase at the operation pH. On the other hand, at high temperature (65 °C), non-specific interactions with the agarose carrier prevailed, evidenced by the presence of myoglobin in the eluate.

Original languageEnglish
Pages (from-to)2401-2407
Number of pages7
JournalAnalytical and bioanalytical chemistry
Volume397
Issue number6
DOIs
Publication statusPublished - Jul 1 2010

Keywords

  • Affinity chromatography
  • Glycoprotein enrichment
  • Temperature control

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry

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