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    Selective immobilization of Sonic hedgehog on benzylguanine terminated patterned self-assembled monolayers

    Year: 2011

    Journal: Biomaterials, Volume 32, Issue 28, October 2011, Pages 6719-6728, 20110906

    Authors: Kwok C-W 1, Strähle U. 1 2, Zhao Y 3, Scharnweber T. 2, Weigel S. 2, Welle A. 2

    Last authors: Alexander Welle

    Organizations: 1 Karlsruhe Institute of Technology, Institute for Toxicology and Genetics, Germany 2 Karlsruhe Institute of Technology, Institute for Biological Interfaces, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany 3 SensoPath Technologies Inc., Bozeman, MT 59718, USA

    Country: Germany, USA, US, United States, United States of America, America

    Patterned two-component, self-assembled monolayers on gold were produced by UV lithography. An oligo(ethylene glycol) terminated disulfide served as inert matrix reducing unspecific protein adsorption and cell adhesion. The second component of the self-assembled monolayer (SAM) presented a benzylguanine moiety for the immobilization of Sonic hedgehog (Shh) fused to a mutant O6-alkylguanine-DNA alkyltransferase (SNAP-tag™). The enzymatic activity of the SNAP-tag allows selective and covalent immobilization of the linked Shh. Time-of-flight secondary ion mass spectrometry verified the correct lateral distribution of the benzylguanine head groups in the patterned SAM. The quantification of unspecific and specific protein binding to mixed SAMs showed increased adsorption of albumin with increasing benzylguanine/(ethylene glycol) ratios. However, the immobilization of SNAP-tagged Shh was not blocked by pre-adsorbed albumin. Furthermore, the obtained micro-patterned substrates permitted direct immobilization of SNAP-tagged Shh even in the presence of many competing proteins from conditioned media of transfected HEK293 cells. Therefore, the presented system is suited for the controlled immobilization of fusion proteins from complex mixtures avoiding purification steps.