Disentangling Glycan-Protein Interactions: Nuclear Magnetic Resonance (NMR) to the Rescue

Sara Bertuzzi; Ana Poveda; Ana Ard&#225;; Ana Gimeno; Jes&#250;s Jim&#233;nez-Barbero

doi:10.3791/66530

JoVE Journal
Chemistry

This content is Free Access.

JoVE Journal Chemistry

Disentangling Glycan-Protein Interactions: Nuclear Magnetic Resonance (NMR) to the Rescue

糖鎖-タンパク質相互作用の解きほぐし:核磁気共鳴(NMR)による解決

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07:40 min

May 17, 2024

DOI: 10.3791/66530-v

Sara Bertuzzi¹, Ana Poveda¹, Ana Ardá^1,2, Ana Gimeno^1,2, Jesús Jiménez-Barbero^1,2,3

¹CIC bioGUNE, Basque Research and Technology Alliance (BRTA), ²lkerbasque, Basque Foundation for Science, ³Centro de Investigacion Biomedica En Red de Enfermedades Respiratorias

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Overview

This protocol details the acquisition, processing, and analysis of NMR experiments to characterize protein-glycan interactions in solution. It outlines methodologies that contribute to structural glycobiology and molecular recognition studies.

Key Study Components

Area of Science

Structural Glycobiology
Molecular Recognition
Biophysical Chemistry

Background

Exploration of glycan lectin systems related to health and disease.
Integration of chemistry, biology, and biomedicine methodologies.
Advancements in glycosciences for glycan-based therapies.
Importance of NMR methodologies in studying glycan interactions.

Purpose of Study

To provide a detailed protocol for NMR experiments.
To elucidate the structural and dynamic aspects of glycan interactions.
To enhance understanding of molecular recognition processes.

Methods Used

Preparation of protein-ligand complex samples.
Utilization of saturation transfer difference and chemical shift perturbation analysis.
Execution of NMR experiments with specific pulse sequences.
Data processing and comparative analysis of NMR spectra.

Main Results

Identification of binding epitopes through STD NMR signals.
Measurement of chemical shift perturbations in protein-ligand interactions.
Construction of 2D plots to visualize binding site interactions.
Documentation of integrals for signal intensities in spectra.

Conclusions

The protocol provides a comprehensive approach to studying glycan interactions.
Findings contribute to the understanding of glycan-based molecular recognition.
Potential implications for developing advanced glycan therapies.

Frequently Asked Questions

What is the main focus of this study?

The study focuses on characterizing protein-glycan interactions using NMR methodologies.

What methodologies are highlighted in the protocol?

The protocol highlights saturation transfer difference and chemical shift perturbation analysis.

How does this research contribute to glycosciences?

It enhances understanding of molecular recognition and supports the development of glycan-based therapies.

What are the key steps in the NMR experiment?

Key steps include sample preparation, setting up the NMR instrument, and data processing.

What results can be expected from the NMR analysis?

Results include identification of binding sites and measurement of chemical shift perturbations.

Is this protocol applicable to other glycan systems?

Yes, the protocol can be adapted for various glycan-lectin interactions.

ここでは、溶液中のタンパク質-糖鎖相互作用の特性評価を目的とした一連のNMR実験の取得、処理、および分析を詳述したプロトコールを紹介します。最も一般的なリガンドベースおよびタンパク質ベースの方法論が概説されており、構造糖鎖生物学および分子認識研究の分野に貢献していることは間違いありません。

私たちの研究では、多様な糖鎖レクチンシステムの研究を掘り下げ、健康や疾患との関連性を探っています。私たちは、化学、生物学、生物医学の方法論を融合させ、がんから細菌やウイルスの感染に至るまで、病理学者に対する免疫応答の役割を深く調査しました。近年、糖鎖科学は、糖鎖の設計、合成、構造分子認識の解析を原子レベルから細胞レベルに変革することで、画期的なマイルストーンを打ち立てています。

これらの進歩は、蛍光療法または先進的糖鎖ベースの治療の舞台を整えるものです。このプロトコールでは、糖鎖分野で採用されている2つの最も強力なNMR方法論を取得、処理、分析するための手順を概説しています。これらの方法論、飽和移動差、および化学的安全性摂動分析は、地衣類の分子認識の構造的および動的側面の正確な情報を補完的に提供します。