Journal
JOURNAL OF BIOSCIENCE AND BIOENGINEERING
Volume 125, Issue 2, Pages 192-198Publisher
SOC BIOSCIENCE BIOENGINEERING JAPAN
DOI: 10.1016/j.jbiosc.2017.09.006
Keywords
Alanine scanning; Atomic force microscopy; Flow-quartz crystal microbalance; Four-helix bundle; Phage display; Metal-binding sequence
Funding
- MEXT-Supported Program for the Strategic Research Foundation at Private Universities [S1512002]
- JSPS KAKENHI [16K14494]
- Grants-in-Aid for Scientific Research [16K14494] Funding Source: KAKEN
Ask authors/readers for more resources
Protein metal hybrids are functional materials with various industrial applications. For example, a redox enzyme immobilized on a platinum electrode is a key component of some biofuel cells and biosensors. To create these hybrid materials, protein molecules are bound to metal surfaces. Here, we report the selection of a novel platinum-binding sequence in a loop of a four-helix bundle protein, the Lac repressor four-helix protein (LARFH), an artificial protein in which four identical alpha-helices are connected via three identical loops. We created a genetic library in which the Ser-Gly-Gln-Gly-Gly-Ser sequence within the first inter-helical loop of LARFH was semi-randomly mutated. The library was then subjected to selection for platinum-binding affinity by using the T7 phage display method. The majority of the selected variants contained the Tyr-Lys-Arg-Gly-Tyr-Lys (YKRGYK) sequence in their randomized segment. We characterized the platinum-binding properties of mutant LARFH by using quartz crystal microbalance analysis. Mutant LARFH seemed to interact with platinum through its loop containing the YKRGYK sequence, as judged by the estimated exclusive area occupied by a single molecule. Furthermore, a 10-residue peptide containing the YKRGYK sequence bound to platinum with reasonably high affinity and basic side chains in the peptide were crucial in mediating this interaction. In conclusion, we have identified an amino acid sequence, YKRGYK, in the loop of a helix-loop-helix motif that shows high platinum-binding affinity. This sequence could be grafted into loops of other polypeptides as an approach to immobilize proteins on platinum electrodes for use as biosensors among other applications. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available