Blue protein with red fluorescence

The walleye (Sander vitreus) is a golden yellow fish that inhabits the Northern American lakes. The recent sightings of the blue walleye and the correlation of its sighting to possible increased UV radiation have been proposed earlier. The underlying molecular basis of its adaptation to increased UV radiation is the presence of a protein (Sandercyanin)–ligand complex in the mucus of walleyes. Degradation of heme by UV radiation results in the formation of Biliverdin IXα (BLA), the chromophore bound to Sandercyanin. We show that Sandercyanin is a monomeric protein that forms stable homotetramers on addition of BLA to the protein. A structure of the Sandercyanin–BLA complex, purified from the fish mucus, reveals a glycosylated protein with a lipocalin fold. This protein–ligand complex absorbs light in the UV region (λmax of 375 nm) and upon excitation at this wavelength emits in the red region (λmax of 675 nm). Unlike all other known biliverdin-bound fluorescent proteins, the chromophore is noncovalently bound to the protein. We provide here a molecular rationale for the observed spectral properties of Sandercyanin.

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Fluorescent protein expression in temperature tolerant and susceptible reef-building corals

Journal of the Marine Biological Association of the United Kingdom ◽

10.1017/s0025315421000059 ◽

2021 ◽

pp. 1-10

Author(s):

Exequiel Gabriel S. Dizon ◽

Jeric P. Da-Anoy ◽

Melissa S. Roth ◽

Cecilia Conaco

Keyword(s):

Spectral Properties ◽

Coral Species ◽

Fluorescent Protein ◽

Fluorescent Proteins ◽

Expression Profiles ◽

Expression Patterns ◽

Antioxidant Properties ◽

Variable Expression ◽

Acropora Digitifera ◽

Insight Into

Abstract Fluorescent proteins (FPs) are reported to play an important role as photoprotectants and antioxidants in corals subjected to stressful conditions. Identifying the various FP genes expressed and FP gene expression patterns under stress in diverse coral species can provide insight into FP function. In this study, we identified 16 putative FP homologues from the transcriptomes of corals with varying susceptibility to elevated temperature, including Acropora digitifera, Favites colemani, Montipora digitata and Seriatopora caliendrum. Each coral expressed a different complement of FP transcripts, which were predicted to have distinct spectral properties. The most diverse and abundant repertoire of FP transcripts, including at least 6 green FPs, were expressed in the temperature-tolerant coral, F. colemani. In comparison, the other corals expressed fewer FP types. Specific FP transcripts exhibited variable expression profiles in coral fragments subjected to 32 ± 1 °C (treatment) or 28 ± 1 °C (control) for up to 72 h, suggesting that distinct FPs may have different roles. Further studies on the expression of the proteins encoded by these FP transcripts, their fluorescence activity, tissue localization, and possible antioxidant properties, are needed to reveal their contribution to thermal stress tolerance in certain species of corals.

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Influence of UV Radiation on the Spectral Properties of 2-METYL-4-Chlorophenoxy Propionic Acids

Russian Physics Journal ◽

10.1007/s11182-020-02187-0 ◽

2020 ◽

Vol 63 (8) ◽

pp. 1424-1428

Author(s):

I. V. Sokolova ◽

A. A. Solokha ◽

O. N. Tchaikovskaya

Keyword(s):

Uv Radiation ◽

Spectral Properties

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UV-induced Hyperphosphorylation of Replication Protein A Depends on DNA Replication and Expression of ATM Protein

Molecular Biology of the Cell ◽

10.1091/mbc.12.5.1199 ◽

2001 ◽

Vol 12 (5) ◽

pp. 1199-1213 ◽

Cited By ~ 58

Author(s):

Gregory G. Oakley ◽

Lisa I. Loberg ◽

Jiaqin Yao ◽

Mary A. Risinger ◽

Remy L. Yunker ◽

...

Keyword(s):

Dna Replication ◽

Uv Radiation ◽

Excision Repair ◽

Genetic Disorder ◽

Protein A ◽

Dna Recombination ◽

Delayed Response ◽

Replication Intermediates

Exposure to DNA-damaging agents triggers signal transduction pathways that are thought to play a role in maintenance of genomic stability. A key protein in the cellular processes of nucleotide excision repair, DNA recombination, and DNA double-strand break repair is the single-stranded DNA binding protein, RPA. We showed previously that the p34 subunit of RPA becomes hyperphosphorylated as a delayed response (4–8 h) to UV radiation (10–30 J/m2). Here we show that UV-induced RPA-p34 hyperphosphorylation depends on expression of ATM, the product of the gene mutated in the human genetic disorder ataxia telangiectasia (A-T). UV-induced RPA-p34 hyperphosphorylation was not observed in A-T cells, but this response was restored by ATM expression. Furthermore, purified ATM kinase phosphorylates the p34 subunit of RPA complex in vitro at many of the same sites that are phosphorylated in vivo after UV radiation. Induction of this DNA damage response was also dependent on DNA replication; inhibition of DNA replication by aphidicolin prevented induction of RPA-p34 hyperphosphorylation by UV radiation. We postulate that this pathway is triggered by the accumulation of aberrant DNA replication intermediates, resulting from DNA replication fork blockage by UV photoproducts. Further, we suggest that RPA-p34 is hyperphosphorylated as a participant in the recombinational postreplication repair of these replication products. Successful resolution of these replication intermediates reduces the accumulation of chromosomal aberrations that would otherwise occur as a consequence of UV radiation.

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The Kindling Fluorescent Protein: A Transient Photoswitchable Marker

Physiology ◽

10.1152/physiol.00056.2005 ◽

2006 ◽

Vol 21 (3) ◽

pp. 162-170 ◽

Cited By ~ 32

Author(s):

J. Nathan Henderson ◽

S. James Remington

Keyword(s):

Fluorescent Protein ◽

Fluorescent Proteins ◽

Protein A ◽

Cellular Imaging ◽

Continuous Observation ◽

Protein Markers ◽

Background Fluorescence ◽

Direct Tracking

Passive fluorescent protein markers are indispensable for dynamic cellular imaging; however, they are unselective, introduce constant background fluorescence, and require continuous observation. Photoactivatable fluorescent proteins have now been developed whose fluorescence can be switched on and off by illumination, allowing selective and direct tracking of tagged objects without the need for continuous imaging. The “kindling fluorescent protein” is a photoactivatable marker with a novel twist: it turns itself off after a selectable period.

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Essential Role for OspA/B in the Life Cycle of the Lyme Disease Spirochete

Journal of Experimental Medicine ◽

10.1084/jem.20031960 ◽

2004 ◽

Vol 199 (5) ◽

pp. 641-648 ◽

Cited By ~ 218

Author(s):

Xiaofeng F. Yang ◽

Utpal Pal ◽

Sophie M. Alani ◽

Erol Fikrig ◽

Michael V. Norgard

Keyword(s):

Life Cycle ◽

Lyme Disease ◽

Outer Surface ◽

Molecular Basis ◽

Potential Role ◽

Virulent Strain ◽

Protein A ◽

Complex Life Cycle ◽

Close Relative ◽

Deficient Mutant

The molecular basis of how Borrelia burgdorferi (Bb), the Lyme disease spirochete, maintains itself in nature via a complex life cycle in ticks and mammals is poorly understood. Outer surface (lipo)protein A (OspA) of Bb has been the most intensively studied of all borrelial molecular constituents, and hence, much has been speculated about the potential role(s) of OspA in the life cycle of Bb. However, the precise function of OspA (along with that of its close relative and operonic partner, outer surface [lipo]protein B [OspB]) heretofore has not been directly determined, due primarily to the inability to generate an OspA/B-deficient mutant from a virulent strain of Bb. In this study, we created an OspA/B-deficient mutant of an infectious human isolate of Bb (strain 297) and found that OspA/B function was not required for either Bb infection of mice or accompanying tissue pathology. However, OspA/B function was essential for Bb colonization of and survival within tick midguts, events crucial for sustaining Bb in its natural enzootic life cycle.

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Orange Fluorescent Proteins: Filling the Spectral Gap

Acta Crystallographica Section A Foundations and Advances ◽

10.1107/s2053273314083296 ◽

2014 ◽

Vol 70 (a1) ◽

pp. C1670-C1670

Author(s):

Sergei Pletnev ◽

Daria Shcherbakova ◽

Oksana Subach ◽

Vladimir Malashkevich ◽

Steven Almo ◽

...

Keyword(s):

In Vivo Imaging ◽

Spectral Properties ◽

Molecular Mechanisms ◽

Fluorescent Protein ◽

Fluorescent Proteins ◽

Stokes Shift ◽

Emission Wavelength ◽

Microscopy Imaging ◽

Large Stokes Shift

Fluorescent proteins (FPs) have become valuable tools for molecular biology, biochemistry, medicine, and cancer research. Starting from parent green fluorescent protein (GFP), most challenging task of the FPs studies was the development of FPs with longer excitation/emission wavelength. This pursuit was motivated by advantages of so-called red-shifted FPs, namely, lower background of cellular autofluorescence in microscopy, lower light scattering and reduced tissue absorbance of longer wavelengths for in vivo imaging. In addition to FPs with regular spectral properties, there are proteins of other types available, including FPs with a large Stokes shift and photoconvertible FPs. These special kinds of FPs have become useful in super-resolution microscopy, imaging of enzyme activities, protein-protein interactions, photolabeling, and in vivo imaging. According to their emission wavelength, red-shifted FPs could be divided in the following groups: 520-540 nm yellow FPs (YFPs), 540-570 nm orange FPs (OFPs), 570-620 nm red FPs (RFPs), and > 620 nm far-RFPs. Red shift of the excitation/emission bands of these FPs is predominantly achieved by extension of the conjugated system of the chromophore and its protonation/deprotonation. The variety of spectral properties of FPs (excitation and emission wavelength, quantum yield, brightness, photo- and pH- stability, photoconversion, large Stokes shift, etc) results from the different chromophore structures and its interactions with surrounding amino acid residues. In this work we focus on structural studies and molecular mechanisms of FPs with orange emission.

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Stalling chromophore maturation of the fluorescent protein Venus reveals the molecular basis of the final oxidation step

10.1101/2020.10.13.337386 ◽

2020 ◽

Author(s):

Husam Sabah Auhim ◽

Bella L. Grigorenko ◽

Tessa Harris ◽

Igor V. Polyakov ◽

Colin Berry ◽

...

Keyword(s):

Molecular Basis ◽

Fluorescent Protein ◽

Fluorescent Proteins ◽

Life Sciences ◽

Intermediate Stage ◽

Open Conformation ◽

Oxidation Step ◽

Canonical Amino Acid

AbstractFluorescent proteins (FPs) have revolutionised the life sciences but the mechanism of chromophore maturation is still not fully understood. Incorporation of a photo-responsive non-canonical amino acid within the chromophore stalls maturation of Venus, a yellow FP, at an intermediate stage; the crystal structure reveals the presence of O2 located above a dehydrated enolate imidazolone (I) ring, close to the strictly conserved Gly67 that occupies a twisted conformation. His148 adopts an “open” conformation, potentially allowing O2 access to the chromophore. Absorption spectroscopy supported by QM/MM simulations suggest that the first oxidation step involves formation of a hydroperoxyl intermediate in conjunction with dehydrogenation of the methylene bridge. A fully conjugated mature chromophore is formed through release of H2O2 upon irradiation of this intermediate, both in vitro and in vivo. The possibility of interrupting and photochemically restarting chromophore maturation, and the mechanistic insights opens up new approaches for engineering optically controlled fluorescent proteins.

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326 GENERATION OF A DOUBLE-TRANSGENIC PIG WITH PANCREAS-SPECIFIC GREEN AND LIVER-SPECIFIC RED FLUORESCENCE

Reproduction Fertility and Development ◽

10.1071/rdv25n1ab326 ◽

2013 ◽

Vol 25 (1) ◽

pp. 311 ◽

Cited By ~ 1

Author(s):

H. Matsunari ◽

K. Nakano ◽

T. Kanai ◽

R. Sakai ◽

M. Watanabe ◽

...

Keyword(s):

Gene Transfer ◽

Fluorescent Proteins ◽

Green Fluorescence ◽

Tissue Specific ◽

Specific Fluorescence ◽

Red Fluorescence ◽

Feasible Option ◽

Organ Tissue ◽

Transfer Method

Transgenic (Tg) pigs with organ/tissue-specific fluorescence expression provide invaluable research tools for many types of studies, such as organogenesis analysis, in vitro tissue generation from pluripotent cells, and progenitor/stem cell transplantation therapy. We aimed to develop a Tg pig characterised by pancreas- and liver-specific fluorescence expression. A 8.4 kb transgene construct expressing Venus (green fluorescence) under the control of the mouse Pdx1 (pancreatic duodenal homeobox-1) promoter and a BAC-derived construct (170 kb) consisting of the whole-length porcine albumin (Alb) promoter and humanized Kusabira-Orange (huKO, red fluorescence) was introduced into porcine in vitro-matured oocytes using the intracytoplasmic sperm injection (ICSI)-mediated gene transfer method. Injected embryos were transferred to the oviducts of oestrus-synchronized recipients after culture for 1 to 3 days. The transfer of 370 Pdx1-Venus embryos into 4 recipients produce 22 (5.9%) fetuses/piglets, and 9 (40.9%) Tg pigs exhibited pancreas-specific Venus expression. Two (1 male and 1 female) founder Pdx1-Venus-Tg pigs were mated with wild-type (WT) pigs and produced 32 offspring in 3 litters, of which 16 (50.0%) were transgenic. Pancreas-specific Venus expression was inherited in these Tg offspring. The transfer of 523 Alb-huKO embryos into 4 recipients resulted in 19 (3.6%) piglets including a Tg female, which showed liver-specific huKO fluorescent expression. Expression of huKO was detected by RT-PCR exclusively in liver, but not in 7 other organs/tissues examined, including heart, lung, stomach, small intestine, spleen, kidney and skin. This founder Tg female produced a total of 12 non-Tg and 5 Tg offspring (in 2 litters) after mating with a WT boar. Liver-specific huKO expression was inherited in these Tg offspring. Furthermore, the mating of a female Pdx1-Venus pig with an Alb-huKO boar yielded 7 non-Tg and 10 Tg pigs. Four of these Tg pigs carrying both of the transgenes exhibited both pancreas-specific Venus and liver-specific huKO expression in single individuals. Double-Tg pigs with pancreas-specific green fluorescence and liver-specific red fluorescence grew normally, and tests of their reproduction ability are currently underway. These data demonstrate that transgene introduction by ICSI-mediated gene transfer into in vitro-matured oocytes is a feasible option for generating pigs expressing fluorescent proteins in a tissue-specific manner.

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