Ru-Porphyrin Protein Scaffolds for Sensing O₂

Abstract

Hemoprotein-based scaffolds containing phosphorescent ruthenium(II) CO mesoporphyrin IX (RuMP) are reported here for oxygen (O2) sensing in biological contexts. RuMP was incorporated into the protein scaffolds during protein expression utilizing a novel method that we have described previously. A high-resolution (2.00 Å) crystal structure revealed that the unnatural porphyrin binds to the proteins in a manner similar to the native heme and does not perturb the protein fold. The protein scaffolds were found to provide unique coordination environments for RuMP and modulate the porphyrin emission properties. Emission lifetime measurements demonstrate a linear O2 response within the physiological range and precision comparable to commercial O2 sensors. The RuMP proteins are robust, readily-modifiable platforms and display promising O2 sensing properties for future in vivo applications.

Tunnels modulate ligand flux in a heme nitric oxide/oxygen binding (H-NOX) domain

“…Ns H-NOX constructs were transformed into the RP523 strain of Escherichia coli (45). Expression cultures were grown at 37°C in Terrific Broth in the presence of 30 μg∕mL hemin (approximately 200× stock in DMSO) and 75-100 μg∕mL ampicillin.…”

“…Proteins were isolated in the Fe II -unligated state (Soret maxima of 429-430 nm). Tt H-NOX constructs were purified as described previously (26,45) and isolated in the Fe II -O 2 ligation state (Soret maximum of approximately 416 nm).…”

Conformational control of cofactors in nature – the influence of protein-induced macrocycle distortion on the biological function of tetrapyrroles

“…We are currently developing an alternative approach that can account for this phenomenon and will use it to assess the conformational effects upon site-energy distribution amongst the BChls in the FMO complex. 244,245 and in particular, for the highly non-planar conformation imposed by the protein on its single heme-cofactor. 246,247,248,249,250,251,252 This is one of the most distorted heme-cofactors to be observed in natural systems, 247 and this feature has been linked to its uncommonly high midpoint potential 248 and is likely crucial to its biological function.…”

“…244 The . This highlights the complicated relationship between the effects of distortion in concert with other environmental aspects.…”

Non-invasive monitoring of skin inflammation using an oxygen-sensing paint-on bandage

“…The relationship between emission intensity and oxygen concentration (pO 2 ) is described by the Stern-Volmer equation, shown below [11]. This phenomenon of oxygen-dependent quenching of phosphorescence has been exploited frequently for various biomedical applications [12][13][14][15][16][17][18][19]. As has been described by Wilson and Vinogradov, this process is highly efficient and responds instantaneously to fluctuations or perturbations in oxygen concentration.…”