A murine monoclonal antibody to glycogen: characterization of epitope-fine specificity by saturation transfer difference (STD) NMR spectroscopy and its use in mycobacterial capsular α-glucan research

Chembiochem. 2015 Apr 13;16(6):977-89. doi: 10.1002/cbic.201402713. Epub 2015 Mar 12.

Abstract

Mycobacterium tuberculosis, the causative agent of tuberculosis (TB), is a major pathogen responsible for 1.5 million deaths annually. This bacterium is characterized by a highly unusual and impermeable cell envelope, which plays a key role in mycobacterial survival and virulence. Although many studies have focused on the composition and functioning of the mycobacterial cell envelope, the capsular α-glucan has received relatively minor attention. Here we show that a murine monoclonal antibody (Mab) directed against glycogen cross-reacts with mycobacterial α-glucans, polymers of α(1-4)-linked glucose residues with α(1-6)-branch points. We identified the Mab epitope specificity by saturation transfer difference NMR and show that the α(1-4)-linked glucose residues are important in glucan-Mab interaction. The minimal epitope is formed by (linear) maltotriose. Notably, a Mycobacterium mutant lacking the branching enzyme GlgB does not react with the Mab; this suggests that the α(1-6)-branches form part of the epitope. These seemingly conflicting data can be explained by the fact that in the mutant the linear form of the α-glucan (amylose) is insoluble. This Mab was subsequently used to develop several techniques helpful in capsular α-glucan research. By using a capsular glucan-screening methodology based on this Mab we were able to identify several unknown genes involved in capsular α-glucan biogenesis. Additionally, we developed two methods for the detection of capsular α-glucan levels. This study therefore opens new ways to study capsular α-glucan and to identify possible targets for further research.

Keywords: STD-NMR spectroscopy; antibodies; carbohydrates; epitope specificity; glycogen monoclonal antibody; mycobacteria.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal / immunology*
  • Antibody Specificity*
  • Bacterial Capsules / metabolism*
  • Cell Wall / metabolism
  • DNA Transposable Elements / genetics
  • Epitopes / immunology*
  • Glycogen / biosynthesis
  • Glycogen / chemistry
  • Glycogen / immunology*
  • Glycogen / metabolism*
  • Magnetic Resonance Spectroscopy
  • Mice
  • Mutation
  • Mycobacterium / cytology
  • Mycobacterium / metabolism*
  • Oligosaccharides / chemistry

Substances

  • Antibodies, Monoclonal
  • DNA Transposable Elements
  • Epitopes
  • Oligosaccharides
  • Glycogen