title: Micromolar biosensing of nitric oxide using myoglobin immobilized in a synthetic silk film.
abstract: In this work we investigate the use of coiled-coil silk proteins, produced in recombinant Escherichia coli, as a new material for immobilizing biosensors. Myoglobin was embedded in transparent honeybee silk protein films. Immobilized myoglobin maintained a high affinity for nitric oxide (KD NO=52 µM) and good sensitivity with a limit of detection of 5 µM. The immobilized myoglobin-silk protein film was stable and could be stored as a dry film at room temperature for at least 60 days. The effect of immobilization on the structure of myoglobin was fully investigated using UV/visible, Fourier Transform Infrared and Raman spectroscopy, which indicated a weakening in the strength of the iron-histidine bond. This study demonstrates that recombinant coiled-coil silk proteins provide a safe and environmentally friendly alternative to sol-gels for stabilizing heme proteins for use as optical biosensors.
authors.affiliation: Ecosystem Sciences, CSIRO, GPO Box 1700, Acton, ACT 2601, Australia; Food Futures Flagship, CSIRO, GPO Box 1700, Acton, ACT 2601, Australia. Electronic address: trevor.rapson@csiro.au.
authors.forename: Trevor D
authors.initials: TD
authors.lastname: Rapson
authors.affiliation: Materials Science and Engineering, CSIRO, PO Box 21, Belmont, VIC 3216, Australia.
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Church
authors.affiliation: Ecosystem Sciences, CSIRO, GPO Box 1700, Acton, ACT 2601, Australia.
authors.forename: Holly E
authors.initials: HE
authors.lastname: Trueman
authors.affiliation: Ecosystem Sciences, CSIRO, GPO Box 1700, Acton, ACT 2601, Australia; Food Futures Flagship, CSIRO, GPO Box 1700, Acton, ACT 2601, Australia.
authors.forename: Helen
authors.initials: H
authors.lastname: Dacres
authors.affiliation: Ecosystem Sciences, CSIRO, GPO Box 1700, Acton, ACT 2601, Australia.
authors.forename: Tara D
authors.initials: TD
authors.lastname: Sutherland
authors.affiliation: Food Futures Flagship, CSIRO, GPO Box 1700, Acton, ACT 2601, Australia.
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Trowell
mesh_terms: D000818:Animals; D001516:Bees; D015374:Biosensing Techniques / Q000379:methods*; D006801:Humans; D055767:Immobilized Proteins / Q000737:chemistry; D009211:Myoglobin / Q000737:chemistry; D009569:Nitric Oxide / Q000032:analysis*; D010084:Oxidation-Reduction; D055550:Protein Stability; D011994:Recombinant Proteins / Q000737:chemistry; D047011:Silk / Q000737:chemistry; D013059:Spectrum Analysis, Raman
chemical_list: D055767:Immobilized Proteins; D009211:Myoglobin; D011994:Recombinant Proteins; D047011:Silk; D009569:Nitric Oxide
keywords: Coiled-coil proteins; Heme proteins; Honeybee silk; Optical biosensors