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

Gronewold, T. M., Baumgartner, A., Hierer, J., Sierra, S., Blind, M., Schäfer, F., et al. (2009). Kinetic binding analysis of aptamers targeting HIV-1 proteins by a combination of a microbalance array and mass spectrometry (MAMS). Journal of proteome research, 8(7), 3568-77. doi:10.1021/pr900265r.

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Item Permalink: https://hdl.handle.net/11858/00-001M-0000-0028-644B-8 Version Permalink: https://hdl.handle.net/11858/00-001M-0000-0028-644C-6
Genre: Journal Article

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 Creators:
Gronewold, T. M., Author
Baumgartner, A., Author
Hierer, J., Author
Sierra, S., Author
Blind, M., Author
Schäfer, F., Author
Blümer, J., Author
Tillmann, T., Author
Kiwitz, A., Author
Kaiser, R., Author
Zabe-Kühn, M., Author
Quandt, E.1, Author
Famulok, M.1, Author
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1External Organizations, ou_persistent22              

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Free keywords: Amino Acid Sequence Aptamers, Peptide/*chemistry Biotinylation HIV Envelope Protein gp120/chemistry HIV Reverse Transcriptase/metabolism HIV-1/*chemistry Humans Kinetics Mass Spectrometry/*methods Molecular Sequence Data Peptides/chemistry Protein Binding Proteins/chemistry Sequence Homology, Amino Acid Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods Viral Proteins/*chemistry
 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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 Dates: 2009
 Publication Status: Issued
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 Identifiers: Other: 19469583
DOI: 10.1021/pr900265r
ISSN: 1535-3893 (Print)
ISSN: 1535-3893 (Linking)
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Title: Journal of proteome research
  Alternative Title : J. Proteome Res.
Source Genre: Journal
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Pages: - Volume / Issue: 8 (7) Sequence Number: - Start / End Page: 3568 - 77 Identifier: -