Identification and solution conformation of multiple epitopes recognized by a MUC2 mucin-specific monoclonal antibody

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Abstract

We have identified the optimal epitope, 21TQTPT25, in the tandem repeat of mucin 2 (MUC2) glycoprotein by using glycoprotein-specific monoclonal antibody, MAb 994, and synthetic, overlapping and truncated oligopeptides corresponding to the sequence 13TPTPTPTGTQTPTT26. We found that peptides containing the 21TQTPT25 sequence were able to inhibit the 994 antibody binding and also peptides 21TQTPT25 and 17TPTGTQTPT25 were the most inhibitory compounds with the lowest IC50 value (IC50=4 and 3μM, respectively) tested. Interestingly, 21TQTPT25 peptide adopts an unordered structure even in TFE, a solvent that promotes an ordered conformation, as detected by circular dichroism and Fourier-transform infrared spectroscopy. However, Thr at position 26 or amidation of Thr25 at the C-terminus results in a much weaker (3 orders of magnitude) MAb interaction, which can be due to the presence of a turn conformation in peptides with a T26 or an amide C-terminus. We have also observed that MAb 994 recognized two other pentapeptides with the TX1TX2T motif, like 13TPTPT17 (IC50=180μM) and 19TGTQP23 (IC50=65μM), whose sequences are present in the native glycoprotein. These findings might suggest that in the MUC2 tandem repeat unit there are multiple antigenic sites available for recognition in underglycosylated tumor tissue and also explain the heteroclitic nature of MAb 994.

Original languageEnglish
Pages (from-to)254-260
Number of pages7
JournalArchives of Biochemistry and Biophysics
Volume410
Issue number2
DOIs
Publication statusPublished - Feb 15 2003

Fingerprint

Mucin-2
Mucins
Inhibitory Concentration 50
Conformations
Epitopes
Monoclonal Antibodies
Glycoproteins
Peptides
Tandem Repeat Sequences
Oligopeptides
Polytetrafluoroethylene
Dichroism
Fourier Transform Infrared Spectroscopy
Circular Dichroism
Amides
Tumors
Tissue
Antibodies
Neoplasms

Keywords

  • Antibody binding
  • Heteroclitic epitope
  • MUC2 mucin
  • Peptide epitope
  • Secondary structure analysis

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

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title = "Identification and solution conformation of multiple epitopes recognized by a MUC2 mucin-specific monoclonal antibody",
abstract = "We have identified the optimal epitope, 21TQTPT25, in the tandem repeat of mucin 2 (MUC2) glycoprotein by using glycoprotein-specific monoclonal antibody, MAb 994, and synthetic, overlapping and truncated oligopeptides corresponding to the sequence 13TPTPTPTGTQTPTT26. We found that peptides containing the 21TQTPT25 sequence were able to inhibit the 994 antibody binding and also peptides 21TQTPT25 and 17TPTGTQTPT25 were the most inhibitory compounds with the lowest IC50 value (IC50=4 and 3μM, respectively) tested. Interestingly, 21TQTPT25 peptide adopts an unordered structure even in TFE, a solvent that promotes an ordered conformation, as detected by circular dichroism and Fourier-transform infrared spectroscopy. However, Thr at position 26 or amidation of Thr25 at the C-terminus results in a much weaker (3 orders of magnitude) MAb interaction, which can be due to the presence of a turn conformation in peptides with a T26 or an amide C-terminus. We have also observed that MAb 994 recognized two other pentapeptides with the TX1TX2T motif, like 13TPTPT17 (IC50=180μM) and 19TGTQP23 (IC50=65μM), whose sequences are present in the native glycoprotein. These findings might suggest that in the MUC2 tandem repeat unit there are multiple antigenic sites available for recognition in underglycosylated tumor tissue and also explain the heteroclitic nature of MAb 994.",
keywords = "Antibody binding, Heteroclitic epitope, MUC2 mucin, Peptide epitope, Secondary structure analysis",
author = "K. Uray and Price, {Michael R.} and Zs. Majer and E. Vass and M. Holl{\'o}si and F. Hudecz",
year = "2003",
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doi = "10.1016/S0003-9861(02)00693-8",
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TY - JOUR

T1 - Identification and solution conformation of multiple epitopes recognized by a MUC2 mucin-specific monoclonal antibody

AU - Uray, K.

AU - Price, Michael R.

AU - Majer, Zs.

AU - Vass, E.

AU - Hollósi, M.

AU - Hudecz, F.

PY - 2003/2/15

Y1 - 2003/2/15

N2 - We have identified the optimal epitope, 21TQTPT25, in the tandem repeat of mucin 2 (MUC2) glycoprotein by using glycoprotein-specific monoclonal antibody, MAb 994, and synthetic, overlapping and truncated oligopeptides corresponding to the sequence 13TPTPTPTGTQTPTT26. We found that peptides containing the 21TQTPT25 sequence were able to inhibit the 994 antibody binding and also peptides 21TQTPT25 and 17TPTGTQTPT25 were the most inhibitory compounds with the lowest IC50 value (IC50=4 and 3μM, respectively) tested. Interestingly, 21TQTPT25 peptide adopts an unordered structure even in TFE, a solvent that promotes an ordered conformation, as detected by circular dichroism and Fourier-transform infrared spectroscopy. However, Thr at position 26 or amidation of Thr25 at the C-terminus results in a much weaker (3 orders of magnitude) MAb interaction, which can be due to the presence of a turn conformation in peptides with a T26 or an amide C-terminus. We have also observed that MAb 994 recognized two other pentapeptides with the TX1TX2T motif, like 13TPTPT17 (IC50=180μM) and 19TGTQP23 (IC50=65μM), whose sequences are present in the native glycoprotein. These findings might suggest that in the MUC2 tandem repeat unit there are multiple antigenic sites available for recognition in underglycosylated tumor tissue and also explain the heteroclitic nature of MAb 994.

AB - We have identified the optimal epitope, 21TQTPT25, in the tandem repeat of mucin 2 (MUC2) glycoprotein by using glycoprotein-specific monoclonal antibody, MAb 994, and synthetic, overlapping and truncated oligopeptides corresponding to the sequence 13TPTPTPTGTQTPTT26. We found that peptides containing the 21TQTPT25 sequence were able to inhibit the 994 antibody binding and also peptides 21TQTPT25 and 17TPTGTQTPT25 were the most inhibitory compounds with the lowest IC50 value (IC50=4 and 3μM, respectively) tested. Interestingly, 21TQTPT25 peptide adopts an unordered structure even in TFE, a solvent that promotes an ordered conformation, as detected by circular dichroism and Fourier-transform infrared spectroscopy. However, Thr at position 26 or amidation of Thr25 at the C-terminus results in a much weaker (3 orders of magnitude) MAb interaction, which can be due to the presence of a turn conformation in peptides with a T26 or an amide C-terminus. We have also observed that MAb 994 recognized two other pentapeptides with the TX1TX2T motif, like 13TPTPT17 (IC50=180μM) and 19TGTQP23 (IC50=65μM), whose sequences are present in the native glycoprotein. These findings might suggest that in the MUC2 tandem repeat unit there are multiple antigenic sites available for recognition in underglycosylated tumor tissue and also explain the heteroclitic nature of MAb 994.

KW - Antibody binding

KW - Heteroclitic epitope

KW - MUC2 mucin

KW - Peptide epitope

KW - Secondary structure analysis

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U2 - 10.1016/S0003-9861(02)00693-8

DO - 10.1016/S0003-9861(02)00693-8

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JO - Archives of Biochemistry and Biophysics

JF - Archives of Biochemistry and Biophysics

SN - 0003-9861

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