Oriented binding of the His(6)-tagged carboxyl-tail of the L-type Ca2+ channel alpha(1)-subunit to a new NTA-functionalized self-assembled monolayer

R Gamsjaeger, B Wimmer, H Kahr, A Tinazli, S Picuric, S Lata, R Tampe, Y Maulet, H J Gruber, P Hinterdorfer, C Romanin

Research output: Contribution to journalArticle (Academic Journal)

27 Citations (Scopus)

Abstract

Oriented stable binding of functional proteins on surfaces is of fundamental interest for receptor/ligand studies in atomic force microscopy (AFM) and surface plasmon resonance (SPR) experiments. Here we have chosen the His(6)-tagged carboxyl-tail (C-tail) of the alpha(1C)-subunit of the L-type Ca2+ channel and calmodulin (CaM) as its cognitive partner as a model system to develop a new functional surface. Covalently attached self-assembled monolayers on ultraflat gold containing NTA-thiols to which the His(6)-tagged C-tail was bound and thiols with triethylene-glycol groups as matrix-thiols represented the system of choice. The topography of this surface was characterized using AFM; its ability to bind C-tail proteins oriented and stable was confirmed by SPR measurements and by complementary force spectroscopy experiments with a CaM4-construct covalently attached to the tip. The developed anchoring strategy can now be used to study receptor/ligand interactions in general applying force spectroscopy and SPR on His(6)-tagged proteins oriented immobilized onto this new NTA-functionalized self-assembled monolayer.

Original languageEnglish
Pages (from-to)5885-5890
Number of pages6
JournalLangmuir
Volume20
Issue number14
DOIs
Publication statusPublished - 6 Jul 2004

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