Glycosylation of recombinant antigenic proteins from Mycobacterium tuberculosis: In silico prediction of protein epitopes and ex vivo biological evaluation of new semi-synthetic glycoconjugates

Teodora Bavaro, Sara Tengattini, Luciano Piubelli, Luciano Piubelli, Francesca Mangione, Roberta Bernardini, Vincenzina Monzillo, Vincenzina Monzillo, Sandra Calarota, Piero Marone, Massimo Amicosante, Loredano Pollegioni, Loredano Pollegioni, Caterina Temporini, Marco Terreni

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Tuberculosis is still one of the most deadly infectious diseases worldwide, and the use of conjugated antigens, obtained by combining antigenic oligosaccharides, such as the lipoarabinomannane (LAM), with antigenic proteins from Mycobacterium tuberculosis (MTB), has been proposed as a new strategy for developing efficient vaccines. In this work, we investigated the effect of the chemical glycosylation on two recombinant MTB proteins produced in E. coli with an additional seven-amino acid tag (recombinant Ag85B and TB10.4). Different semi-synthetic glycoconjugated derivatives were prepared, starting from mannose and two disaccharide analogs. The glycans were activated at the anomeric position with a thiocyanomethyl group, as required for protein glycosylation by selective reaction with lysines. The glycosylation sites and the ex vivo evaluation of the immunogenic activity of the different neo-glycoproteins were investigated. Glycosylation does not modify the immunological activity of the TB10.4 protein. Similarly, Ag85B maintains its B-cell activity after glycosylation while showing a significant reduction in the T-cell response. The results were correlated with the putative B- and T-cell epitopes, predicted using a combination of in silico systems. In the recombinant TB10.4, the unique lysine is not included in any T-cell epitope. Lys30 of Ag85B, identified as the main glycosylation site, proved to be the most important site involved in the formation of T-cell epitopes, reasonably explaining why its glycosylation strongly influenced the T-cell activity. Furthermore, additional lysines included in different epitopes (Lys103, -123 and -282) are also glycosylated. In contrast, B-cell epitopic lysines of Ag85B were found to be poorly glycosylated and, thus, the antibody interaction of Ag85B was only marginally affected after coupling with mono- or disaccharides.

Original languageEnglish
Article number1081
JournalMolecules
Volume22
Issue number7
DOIs
Publication statusPublished - Jul 1 2017

Fingerprint

Glycosylation
Recombinant proteins
Glycoconjugates
Epitopes
Lysine
T-Lymphocyte Epitopes
Proteins
T-cells
Disaccharides
Cells
B-Lymphocyte Epitopes
Monosaccharides
Mannose
Oligosaccharides
Escherichia coli
Polysaccharides
Glycoproteins
Vaccines
Derivatives
Antigens

Keywords

  • Epitope
  • Glycoconjugate vaccines
  • MTB recombinant antigens
  • Neo-glycoproteins

ASJC Scopus subject areas

  • Organic Chemistry

Cite this

Glycosylation of recombinant antigenic proteins from Mycobacterium tuberculosis : In silico prediction of protein epitopes and ex vivo biological evaluation of new semi-synthetic glycoconjugates. / Bavaro, Teodora; Tengattini, Sara; Piubelli, Luciano; Piubelli, Luciano; Mangione, Francesca; Bernardini, Roberta; Monzillo, Vincenzina; Monzillo, Vincenzina; Calarota, Sandra; Marone, Piero; Amicosante, Massimo; Pollegioni, Loredano; Pollegioni, Loredano; Temporini, Caterina; Terreni, Marco.

In: Molecules, Vol. 22, No. 7, 1081, 01.07.2017.

Research output: Contribution to journalArticle

Bavaro, Teodora ; Tengattini, Sara ; Piubelli, Luciano ; Piubelli, Luciano ; Mangione, Francesca ; Bernardini, Roberta ; Monzillo, Vincenzina ; Monzillo, Vincenzina ; Calarota, Sandra ; Marone, Piero ; Amicosante, Massimo ; Pollegioni, Loredano ; Pollegioni, Loredano ; Temporini, Caterina ; Terreni, Marco. / Glycosylation of recombinant antigenic proteins from Mycobacterium tuberculosis : In silico prediction of protein epitopes and ex vivo biological evaluation of new semi-synthetic glycoconjugates. In: Molecules. 2017 ; Vol. 22, No. 7.
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AU - Calarota, Sandra

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