Kinetic binding analysis of aptamers targeting HIV-1 proteins by a combination of a microbalance array and mass spectrometry (MAMS).

2009: ABaumgartner; MBlind; JBlümer; MFamulok; TMGronewold; JHierer; RKaiser; AKiwitz; EQuandt; FSchafer; SSierra; TTillmann; MZabe-Kühn;

J Proteome Res.2009;8(7):3568-77.10.1021/pr900265r.

NLM PMID: 19469583

Article abstract

An enhanced chip-based detection platform was developed by integrating a surface acoustic wave biosensor of the Love-wave type with protein identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF MS). The system was applied to characterize the interaction of aptamers with their cognate HIV-1 proteins. The aptamers, which target two proteins of HIV-1, were identified using an automated in vitro selection platform. For aptamers S66A-C6 and S68B-C5, which target the V3 loop of the HIV-1 envelope protein gp120, KD values of 406 and 791 nM, respectively, were measured. Aptamer S69A-C15 was shown to bind HIV-1 reverse transcriptase (HIV-1 RT) with a KD value of 637 nM when immobilized on the biosensor surface. HIV-1 RT was identified with high significance using MALDI-ToF MS even in crude protein mixtures. The V3-loop of gp120 could be directly identified when using chip-bound purified protein samples. From crude protein mixtures, it could be identified indirectly with high significance via its fusion-partner glutathione-S-transferase (GST). Our data show that the combination of the selectivity of aptamers with a sensitive detection by MS enables the reliable and quantitative analysis of kinetic binding events of protein solutions in real time.

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Last MEDLINE®/PubMed® update: 1st of December 2015