Peptide tags for enhanced cellular and protein adhesion to single-crystalline sapphire

• Published in: Biotechnology and bioengineering 2007-08, Vol.97 (5), p.1009-1020
• Main Authors: Krauland, Eric M., Peelle, Beau R., Wittrup, K. Dane, Belcher, Angela M.
• Format: Article
• Language: English
• Subjects: affinity; Affinity Labels - chemistry; Aluminum Oxide - chemistry; Binding Sites; Biological and medical sciences; Biotechnology; Cell Adhesion - physiology; Cell Culture Techniques - methods; Coated Materials, Biocompatible - chemistry; electrostatic; Fundamental and applied biological sciences. Psychology; lysine; metal oxides; Peptide Library; peptides; Peptides - chemistry; Protein Binding; Saccharomyces cerevisiae - physiology; Saccharomyces cerevisiae Proteins - chemistry; Saccharomyces cerevisiae Proteins - metabolism; yeast display
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.21341

In order to facilitate a novel means for coupling proteins to metal oxides, peptides were identified from a dodecamer peptide yeast surface display library that bound a model metal oxide material, the C, A, and R crystalline faces of synthetic sapphire (α‐Al2O3). Seven rounds of screening yielded peptides enriched in basic amino acids compared to the naive library. While the C‐face had a high background of endogenous yeast cell binding, the A‐ and R faces displayed clear peptide‐mediated cell adhesion. Cell detachment assays showed that cell adhesion strength correlated positively with increasing basicity of expressed peptides. Cell adhesion was also shown to be sensitive to buffer ionic strength as well as incubation with soluble peptide (with half maximal inhibition of cell binding at ∼5 µM peptide). Next, dodecamer peptides cloned into yeast showed that lysine led to stronger interactions than arginine, and that charge distribution affected adhesion strength. We postulate binding to arise from peptide geometries that permit conformation alignment of the basic amino acids towards the surface so that the charged groups can undergo local electrostatic interactions with the surface oxide. Lastly, peptide K1 (‐(GK)6) was cloned onto the c‐terminus of maltose binding protein (MBP) and the resultant mutant protein showed a half‐maximal binding at ∼10−7–10−6 M, which marked a ∼500‐ to 1,000‐fold binding improvement to sapphire's A‐face as compared with wild‐type MBP. Targeting proteins to metal oxide surfaces with peptide tags may provide a facile one‐step alternative coupling chemistry for the formation of protein bioassays and biosensors. Biotechnol. Bioeng. 2007; 97: 1009–1020. © 2007 Wiley Periodicals, Inc.