Multitechnique characterization of adsorbed peptide and protein orientation: LK310 and Protein G B1

Baio, J. E.; Weidner, T.; Samuel, N. T.; McCrea, Keith; Baugh, Loren; Stayton, Patrick S.; Castner, David G.

doi:doi:10.1116/1.3456176

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Journal of Vacuum Science & Technology B / Volume 28 / Issue 4 / PAPERS FROM THE 37th ANNUAL CONFERENCE ON THE PHYSICS AND CHEMISTRY OF SEMICONDUCTOR INTERFACES / Biological interfaces

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J. Vac. Sci. Technol. B 28, C5D1 (2010); http://dx.doi.org/10.1116/1.3456176 (8 pages)

Multitechnique characterization of adsorbed peptide and protein orientation: LK3₁₀ and Protein G B1

J. E. Baio¹, T. Weidner², N. T. Samuel¹, Keith McCrea³, Loren Baugh², Patrick S. Stayton², and David G. Castner⁴

¹Department of Chemical Engineering, National ESCA and Surface Analysis Center for Biomedical Problems, University of Washington, Seattle, Washington 98195
²Department of Bioengineering, National ESCA and Surface Analysis Center for Biomedical Problems, University of Washington, Seattle, Washington 98195
³Emergence Venture Partners, LLC, Berkeley, California 94710
⁴Department of Chemical Engineering, Department of Bioengineering, and National ESCA and Surface Analysis Center for Biomedical Problems, University of Washington, Seattle, Washington 98195

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(Published online 6 July 2010)

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The ability to orient biologically active proteins on surfaces is a major challenge in the design, construction, and successful deployment of many medical technologies. As methods to orient biomolecules are developed, it is also essential to develop techniques that can accurately determine the orientation and structure of these materials. In this study, two model protein and peptide systems are presented to highlight the strengths of three surface analysis techniques for characterizing protein films: time-of-flight secondary-ion mass spectrometry (ToF-SIMS), sum-frequency generation (SFG) vibrational spectroscopy, and near-edge x-ray absorption fine structure (NEXAFS) spectroscopy. First, the orientation of Protein G B1, a rigid 6 kDa domain covalently attached to a maleimide-functionalized self-assembled monolayer, was examined using ToF-SIMS. Although the thickness of the Protein G layer was similar to the ToF-SIMS sampling depth, orientation of Protein G was successfully determined by analyzing the C₂H₅S⁺ intensity, a secondary-ion derived from a methionine residue located at one end of the protein. Next, the secondary structure of a 13-mer leucine-lysine peptide (LK3₁₀) adsorbed onto hydrophilic quartz and hydrophobic fluorocarbon surfaces was examined. SFG spectra indicated that the peptide’s lysine side chains were ordered on the quartz surface, while the peptide’s leucine side chains were ordered on the fluorocarbon surface. NEXAFS results provided complementary information about the structure of the LK3₁₀ film and the orientations of amide bonds within the LK3₁₀ peptide.

ACKNOWLEDGMENTS

NIH Grant Nos. GM-074511 and EB-002027 (NESAC-BIO) provided funding for this study. T.W. thanks the Deutsche Forschungsgemeinschaft for a research fellowship. The authors also thank Daniel Fischer (NIST) and Cherno Jaye (Hunter College) for providing us with the experimental equipment for NEXAFS spectroscopy and their help at the synchrotron. NEXAFS studies were performed at the NSLS, Brookhaven National Laboratory, which is supported by the U.S. Department of Energy, Division of Materials Science and Division of Chemical Sciences.

Article Outline

INTRODUCTION
EXPERIMENT
1. Protein G B1 films
2. ToF-SIMS
3. Peptide films
4. NEXAFS spectroscopy
5. SFG spectroscopy
RESULTS AND DISCUSSION
1. Probing Protein G B1 orientation with ToF-SIMS
2. Probing the orientation and secondary structure of 3₁₀ -helical LK peptides with SFG and NEXAFS spectroscopy.
CONCLUSIONS

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KEYWORDS and PACS

Keywords

adsorption, EXAFS, hydrophilicity, hydrophobicity, molecular biophysics, monolayers, optical frequency conversion, proteins, secondary ion mass spectra, self-assembly, time of flight mass spectra, XANES, Protein G B1, leucine-lysine peptide, maleimide-oligo(ethylene glycol) self-assembled monolayers, hexafluoropropylene

PACS

87.15.B-

Structure of biomolecules
87.15.M-

Spectra of biomolecules

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History

Received 25 February 2010
Accepted 1 June 2010
Published online 6 July 2010

Digital Object Identifier

http://dx.doi.org/10.1116/1.3456176

PUBLICATION DATA

ISSN

1071-1023 (print)

1520-8567 (online)

Publisher

American Vacuum Society

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