Relevance of the Poly(ethylene glycol) Linkers in Peptide Surfaces for Proteases Assays

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• 作者：Roza Trzcinska ; Katarzyna Balin ; Jerzy Kubacki ; Magdalena E. Marzec ; Roman Pedrys ; Jacek Szade ; Jerzy Silberring ; Andrzej Dworak ; Barbara Trzebicka
• 刊名：Langmuir
• 出版年：2014
• 出版时间：May 6, 2014
• 年：2014
• 卷：30
• 期：17
• 页码：5015-5025
• 全文大小：583K
• 年卷期：v.30,no.17(May 6, 2014)
• ISSN：1520-5827

文摘

Poly(ethylene glycol)s (PEGs) with different lengths were used as linkers during the preparation of peptide surfaces for protease detection. In the first approach, the PEG monolayers were prepared using a 鈥済rafting to鈥?method on 3-aminopropyltrietoxysilane (APTES)-modified silicon wafers. Protected peptides with a fluorescent marker were synthesized by Fmoc solid phase synthesis. The protected peptide structures enabled their site-specific immobilization onto the PEG surfaces. Alternatively, the PEG-peptide surface was obtained by immobilizing a PEG-peptide conjugate directly onto the modified silicon wafer. The surfaces (composition, grafting density, hydrophilicity, and roughness) were characterized by time-of-flight-secondary ion mass spectrometry (ToF-SIMS), X-ray photoelectron spectroscopy (XPS), contact angle (CA), and atomic force microscopy (AFM). Introducing the PEG linker between the peptide and surface increased their resistance toward nonspecific protein adsorption. The peptide surfaces were examined as analytical platforms to study the action of trypsin as a representative protease. The products of the enzymatic hydrolysis were analyzed by fluorescence spectroscopy, electrospray ionization鈥搈ass spectrometry (ESI-MS), and ToF-SIMS. Conclusions about the optimal length of the PEG linker for the analytical application of PEG-peptide surfaces were drawn. This work demonstrates an effective synthetic procedure to obtain PEG-peptide surfaces as attractive platforms for the development of peptide microarrays.