scholarly journals Genetically encoded phenyl azide photochemistry drives positive and negative functional modulation of a red fluorescent protein

RSC Advances ◽  
2015 ◽  
Vol 5 (95) ◽  
pp. 77734-77738 ◽  
Author(s):  
Samuel C. Reddington ◽  
Sarunas Driezis ◽  
Andrew M. Hartley ◽  
Peter D. Watson ◽  
Pierre J. Rizkallah ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Red Fluorescent Protein ◽  
Phenyl Azide ◽  
Functional Modulation

Genetically encoded incorporation of phenyl azide chemistry into the autofluorescent protein mCherry can be used to switch on or off fluorescence.

Fluorescent proteins for in vivo imaging, where's the biliverdin?

2020 ◽  
Vol 48 (6) ◽  
pp. 2657-2667
Author(s):  
Felipe Montecinos-Franjola ◽  
John Y. Lin ◽  
Erik A. Rodriguez
Keyword(s):  
Hydrogen Peroxide ◽  
In Vivo Imaging ◽  
Near Infrared ◽  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Fluorescent Imaging ◽  
Infrared Light ◽  
Red Fluorescent Protein ◽  
Color Palette

Noninvasive fluorescent imaging requires far-red and near-infrared fluorescent proteins for deeper imaging. Near-infrared light penetrates biological tissue with blood vessels due to low absorbance, scattering, and reflection of light and has a greater signal-to-noise due to less autofluorescence. Far-red and near-infrared fluorescent proteins absorb light >600 nm to expand the color palette for imaging multiple biosensors and noninvasive in vivo imaging. The ideal fluorescent proteins are bright, photobleach minimally, express well in the desired cells, do not oligomerize, and generate or incorporate exogenous fluorophores efficiently. Coral-derived red fluorescent proteins require oxygen for fluorophore formation and release two hydrogen peroxide molecules. New fluorescent proteins based on phytochrome and phycobiliproteins use biliverdin IXα as fluorophores, do not require oxygen for maturation to image anaerobic organisms and tumor core, and do not generate hydrogen peroxide. The small Ultra-Red Fluorescent Protein (smURFP) was evolved from a cyanobacterial phycobiliprotein to covalently attach biliverdin as an exogenous fluorophore. The small Ultra-Red Fluorescent Protein is biophysically as bright as the enhanced green fluorescent protein, is exceptionally photostable, used for biosensor development, and visible in living mice. Novel applications of smURFP include in vitro protein diagnostics with attomolar (10−18 M) sensitivity, encapsulation in viral particles, and fluorescent protein nanoparticles. However, the availability of biliverdin limits the fluorescence of biliverdin-attaching fluorescent proteins; hence, extra biliverdin is needed to enhance brightness. New methods for improved biliverdin bioavailability are necessary to develop improved bright far-red and near-infrared fluorescent proteins for noninvasive imaging in vivo.

Site-Specific Protein Covalent Attachment to Nanotubes and Its Electronic Impact on Single Molecule Function

2019 ◽  
Author(s):  
Adam Beachey ◽  
Harley Worthy ◽  
William David Jamieson ◽  
Suzanne Thomas ◽  
Benjamin Bowen ◽  
...  
Keyword(s):  
Single Molecule ◽  
Fluorescent Protein ◽  
Functional Integration ◽  
Attachment Site ◽  
Covalent Attachment ◽  
Great Promise ◽  
Functional Coupling ◽  
Phenyl Azide ◽  
Electronic Impact ◽  
Protein Attachment

<p>Functional integration of proteins with carbon-based nanomaterials such as nanotubes holds great promise in emerging electronic and optoelectronic applications. Control over protein attachment poses a major challenge for consistent and useful device fabrication, especially when utilizing single/few molecule properties. Here, we exploit genetically encoded phenyl azide photochemistry to define the direct covalent attachment of three different proteins, including the fluorescent protein GFP, to carbon nanotube side walls. Single molecule fluorescence revealed that on attachment to SWCNTs GFP’s fluorescence changed in terms of intensity and improved resistance to photobleaching; essentially GFP is fluorescent for much longer on attachment. The site of attachment proved important in terms of electronic impact on GFP function, with the attachment site furthest from the functional center having the larger effect on fluorescence. Our approach provides a versatile and general method for generating intimate protein-CNT hybrid bioconjugates. It can be potentially applied easily to any protein of choice; attachment position and thus interface characteristics with the CNT can easily be changed by simply placing the phenyl azide chemistry at different residues by gene mutagenesis. Thus, our approach will allow consistent construction and modulate functional coupling through changing the protein attachment position.</p>

scholarly journals Novel Regeneration Approach for Creating Reusable FO-SPR Probes with NTA Surface Chemistry

Nanomaterials ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 186
Author(s):  
Jia-Huan Qu ◽  
Karen Leirs ◽  
Remei Escudero ◽  
Žiga Strmšek ◽  
Roman Jerala ◽  
...  
Keyword(s):  
Surface Chemistry ◽  
Fluorescent Protein ◽  
Cost Effective ◽  
Nitrilotriacetic Acid ◽  
Antibody Fragments ◽  
Rapid Screening ◽  
Red Fluorescent Protein ◽  
Effective Manner ◽  
Surface Regeneration ◽  
Sensitivity Specificity

To date, surface plasmon resonance (SPR) biosensors have been exploited in numerous different contexts while continuously pushing boundaries in terms of improved sensitivity, specificity, portability and reusability. The latter has attracted attention as a viable alternative to disposable biosensors, also offering prospects for rapid screening of biomolecules or biomolecular interactions. In this context here, we developed an approach to successfully regenerate a fiber-optic (FO)-SPR surface when utilizing cobalt (II)-nitrilotriacetic acid (NTA) surface chemistry. To achieve this, we tested multiple regeneration conditions that can disrupt the NTA chelate on a surface fully saturated with His6-tagged antibody fragments (scFv-33H1F7) over ten regeneration cycles. The best surface regeneration was obtained when combining 100 mM EDTA, 500 mM imidazole and 0.5% SDS at pH 8.0 for 1 min with shaking at 150 rpm followed by washing with 0.5 M NaOH for 3 min. The true versatility of the established approach was proven by regenerating the NTA surface for ten cycles with three other model system bioreceptors, different in their size and structure: His6-tagged SARS-CoV-2 spike fragment (receptor binding domain, RBD), a red fluorescent protein (RFP) and protein origami carrying 4 RFPs (Tet12SN-RRRR). Enabling the removal of His6-tagged bioreceptors from NTA surfaces in a fast and cost-effective manner can have broad applications, spanning from the development of biosensors and various biopharmaceutical analyses to the synthesis of novel biomaterials.

A brilliant monomeric red fluorescent protein to visualize cytoskeleton dynamics inDictyostelium

FEBS Letters ◽  
2004 ◽  
Vol 577 (1-2) ◽  
pp. 227-232 ◽  
Author(s):  
Markus Fischer ◽  
Ilka Haase ◽  
Evelyn Simmeth ◽  
Günther Gerisch ◽  
Annette Müller-Taubenberger
Keyword(s):  
Fluorescent Protein ◽  
Red Fluorescent Protein ◽  
Cytoskeleton Dynamics

scholarly journals Unique interactions between the chromophore and glutamate 16 lead to far-red emission in a red fluorescent protein

2009 ◽  
Vol 18 (2) ◽  
pp. 460-466 ◽  
Author(s):  
Xiaokun Shu ◽  
Lei Wang ◽  
Leslie Colip ◽  
Karen Kallio ◽  
S. James Remington
Keyword(s):  
Fluorescent Protein ◽  
Red Fluorescent Protein ◽  
Red Emission
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