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Darrin T Schultz - One of the best experts on this subject based on the ideXlab platform.
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laboratory culture of the california sea firefly vargula tsujii ostracoda cypridinidae developing a model system for the evolution of marine bioluminescence
Scientific Reports, 2020Co-Authors: Jessica A Goodheart, Geetanjali Minsky, Mira N Brynjegardbialik, Michael S Drummond, David J Munoz, Timothy R Fallon, Darrin T SchultzAbstract:Bioluminescence, or the production of light by living organisms via chemical reaction, is widespread across Metazoa. Laboratory culture of bioluminescent organisms from diverse taxonomic groups is important for determining the biosynthetic pathways of bioluminescent substrates, which may lead to new tools for biotechnology and biomedicine. Some bioluminescent groups may be cultured, including some cnidarians, ctenophores, and brittle stars, but those use luminescent substrates (Luciferins) obtained from their diets, and therefore are not informative for determination of the biosynthetic pathways of the Luciferins. Other groups, including terrestrial fireflies, do synthesize their own Luciferin, but culturing them is difficult and the biosynthetic pathway for firefly Luciferin remains unclear. An additional independent origin of endogenous bioluminescence is found within ostracods from the family Cypridinidae, which use their luminescence for defense and, in Caribbean species, for courtship displays. Here, we report the first complete life cycle of a luminous ostracod (Vargula tsujii Kornicker & Baker, 1977, the California Sea Firefly) in the laboratory. We also describe the late-stage embryogenesis of Vargula tsujii and discuss the size classes of instar development. We find embryogenesis in V. tsujii ranges from 25-38 days, and this species appears to have five instar stages, consistent with ontogeny in other cypridinid lineages. We estimate a complete life cycle at 3-4 months. We also present the first complete mitochondrial genome for Vargula tsujii. Bringing a luminous ostracod into laboratory culture sets the stage for many potential avenues of study, including learning the biosynthetic pathway of cypridinid Luciferin and genomic manipulation of an autogenic bioluminescent system.
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laboratory culture of the california sea firefly vargula tsujii ostracoda cypridinidae developing a model system for the evolution of marine bioluminescence
bioRxiv, 2019Co-Authors: Jessica A Goodheart, Geetanjali Minsky, Mira N Brynjegardbialik, Michael S Drummond, David J Munoz, Timothy R Fallon, Darrin T Schultz, Jingke Weng, Elizabeth TorresAbstract:Bioluminescence, or the production of light by living organisms via chemical reaction, is widespread across Metazoa. Culturing bioluminescent organisms from diverse taxonomic groups is important for determining the biosynthetic pathways of bioluminescent substrates, which may lead to new tools for biotechnology and biomedicine. Although some previously cultured luminescent groups represent independent origins of bioluminescence, cnidarians, ctenophores, and brittle stars use luminescent substrates (Luciferins) obtained from their diets, and therefore are not informative for determination of the biosynthethic pathways of the Luciferins. Terrestrial fireflies do synthesize their own Luciferin, but the biosynthetic pathway for firefly Luciferin remains unclear. An additional independent origin of endogenous bioluminescence is found within ostracods from the family Cypridinidae, which use their luminescence for defense and, in Caribbean species, for courtship displays. Here, we report the first complete life cycle of a luminous ostracod (Vargula tsujii, the California Sea Firefly) in the laboratory. We also describe the embryonic development of Vargula tsujii and discuss the size classes of instar development. We find embryogenesis in V. tsujii ranges between 25-38 days, and this species appears to have five instar stages, consistent with ontogeny in other cypridinid lineages. We estimate a complete life cycle at 3-4 months. We present the first complete mitochondrial genome for Vargula tsujii. Finally, we find no evidence of significant population genetic structure or cryptic species throughout the southern California range of V. tsujii. Bringing a luminous ostracod into laboratory culture sets the stage for many potential avenues of study, including the biosynthetic pathway of cypridinid Luciferin and genomic manipulation of an autogenic bioluminescent system.
Timothy R Fallon - One of the best experts on this subject based on the ideXlab platform.
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laboratory culture of the california sea firefly vargula tsujii ostracoda cypridinidae developing a model system for the evolution of marine bioluminescence
Scientific Reports, 2020Co-Authors: Jessica A Goodheart, Geetanjali Minsky, Mira N Brynjegardbialik, Michael S Drummond, David J Munoz, Timothy R Fallon, Darrin T SchultzAbstract:Bioluminescence, or the production of light by living organisms via chemical reaction, is widespread across Metazoa. Laboratory culture of bioluminescent organisms from diverse taxonomic groups is important for determining the biosynthetic pathways of bioluminescent substrates, which may lead to new tools for biotechnology and biomedicine. Some bioluminescent groups may be cultured, including some cnidarians, ctenophores, and brittle stars, but those use luminescent substrates (Luciferins) obtained from their diets, and therefore are not informative for determination of the biosynthetic pathways of the Luciferins. Other groups, including terrestrial fireflies, do synthesize their own Luciferin, but culturing them is difficult and the biosynthetic pathway for firefly Luciferin remains unclear. An additional independent origin of endogenous bioluminescence is found within ostracods from the family Cypridinidae, which use their luminescence for defense and, in Caribbean species, for courtship displays. Here, we report the first complete life cycle of a luminous ostracod (Vargula tsujii Kornicker & Baker, 1977, the California Sea Firefly) in the laboratory. We also describe the late-stage embryogenesis of Vargula tsujii and discuss the size classes of instar development. We find embryogenesis in V. tsujii ranges from 25-38 days, and this species appears to have five instar stages, consistent with ontogeny in other cypridinid lineages. We estimate a complete life cycle at 3-4 months. We also present the first complete mitochondrial genome for Vargula tsujii. Bringing a luminous ostracod into laboratory culture sets the stage for many potential avenues of study, including learning the biosynthetic pathway of cypridinid Luciferin and genomic manipulation of an autogenic bioluminescent system.
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laboratory culture of the california sea firefly vargula tsujii ostracoda cypridinidae developing a model system for the evolution of marine bioluminescence
bioRxiv, 2019Co-Authors: Jessica A Goodheart, Geetanjali Minsky, Mira N Brynjegardbialik, Michael S Drummond, David J Munoz, Timothy R Fallon, Darrin T Schultz, Jingke Weng, Elizabeth TorresAbstract:Bioluminescence, or the production of light by living organisms via chemical reaction, is widespread across Metazoa. Culturing bioluminescent organisms from diverse taxonomic groups is important for determining the biosynthetic pathways of bioluminescent substrates, which may lead to new tools for biotechnology and biomedicine. Although some previously cultured luminescent groups represent independent origins of bioluminescence, cnidarians, ctenophores, and brittle stars use luminescent substrates (Luciferins) obtained from their diets, and therefore are not informative for determination of the biosynthethic pathways of the Luciferins. Terrestrial fireflies do synthesize their own Luciferin, but the biosynthetic pathway for firefly Luciferin remains unclear. An additional independent origin of endogenous bioluminescence is found within ostracods from the family Cypridinidae, which use their luminescence for defense and, in Caribbean species, for courtship displays. Here, we report the first complete life cycle of a luminous ostracod (Vargula tsujii, the California Sea Firefly) in the laboratory. We also describe the embryonic development of Vargula tsujii and discuss the size classes of instar development. We find embryogenesis in V. tsujii ranges between 25-38 days, and this species appears to have five instar stages, consistent with ontogeny in other cypridinid lineages. We estimate a complete life cycle at 3-4 months. We present the first complete mitochondrial genome for Vargula tsujii. Finally, we find no evidence of significant population genetic structure or cryptic species throughout the southern California range of V. tsujii. Bringing a luminous ostracod into laboratory culture sets the stage for many potential avenues of study, including the biosynthetic pathway of cypridinid Luciferin and genomic manipulation of an autogenic bioluminescent system.
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SulfoLuciferin is Biosynthesized by a Specialized Luciferin Sulfotransferase in Fireflies.
Biochemistry, 2016Co-Authors: Timothy R Fallon, Maria A. Vicent, Jingke WengAbstract:Firefly Luciferin is a specialized metabolite restricted to fireflies (family Lampyridae) and other select families of beetles (order Coleoptera). Firefly Luciferin undergoes luciferase-catalyzed oxidation to produce light, thereby enabling the luminous mating signals essential for reproductive success in most bioluminescent beetles. Although firefly Luciferin and luciferase have become widely used biotechnological tools, questions remain regarding the physiology and biochemistry of firefly bioluminescence. Here we report sulfoLuciferin to be an in vivo derivative of firefly Luciferin in fireflies and report the cloning of Luciferin sulfotransferase (LST) from the North American firefly Photinus pyralis. LST catalyzes the production of sulfoLuciferin from firefly Luciferin and the sulfo-donor PAPS. SulfoLuciferin is abundant in several surveyed firefly genera as well as in the bioluminescent elaterid beetle Pyrophorus luminosus at a low level. We propose that sulfoLuciferin could serve as a Luciferin storage molecule in fireflies and that LST may find use as a new tool to modulate existing biotechnological applications of the firefly bioluminescent system.
Jessica A Goodheart - One of the best experts on this subject based on the ideXlab platform.
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laboratory culture of the california sea firefly vargula tsujii ostracoda cypridinidae developing a model system for the evolution of marine bioluminescence
Scientific Reports, 2020Co-Authors: Jessica A Goodheart, Geetanjali Minsky, Mira N Brynjegardbialik, Michael S Drummond, David J Munoz, Timothy R Fallon, Darrin T SchultzAbstract:Bioluminescence, or the production of light by living organisms via chemical reaction, is widespread across Metazoa. Laboratory culture of bioluminescent organisms from diverse taxonomic groups is important for determining the biosynthetic pathways of bioluminescent substrates, which may lead to new tools for biotechnology and biomedicine. Some bioluminescent groups may be cultured, including some cnidarians, ctenophores, and brittle stars, but those use luminescent substrates (Luciferins) obtained from their diets, and therefore are not informative for determination of the biosynthetic pathways of the Luciferins. Other groups, including terrestrial fireflies, do synthesize their own Luciferin, but culturing them is difficult and the biosynthetic pathway for firefly Luciferin remains unclear. An additional independent origin of endogenous bioluminescence is found within ostracods from the family Cypridinidae, which use their luminescence for defense and, in Caribbean species, for courtship displays. Here, we report the first complete life cycle of a luminous ostracod (Vargula tsujii Kornicker & Baker, 1977, the California Sea Firefly) in the laboratory. We also describe the late-stage embryogenesis of Vargula tsujii and discuss the size classes of instar development. We find embryogenesis in V. tsujii ranges from 25-38 days, and this species appears to have five instar stages, consistent with ontogeny in other cypridinid lineages. We estimate a complete life cycle at 3-4 months. We also present the first complete mitochondrial genome for Vargula tsujii. Bringing a luminous ostracod into laboratory culture sets the stage for many potential avenues of study, including learning the biosynthetic pathway of cypridinid Luciferin and genomic manipulation of an autogenic bioluminescent system.
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laboratory culture of the california sea firefly vargula tsujii ostracoda cypridinidae developing a model system for the evolution of marine bioluminescence
bioRxiv, 2019Co-Authors: Jessica A Goodheart, Geetanjali Minsky, Mira N Brynjegardbialik, Michael S Drummond, David J Munoz, Timothy R Fallon, Darrin T Schultz, Jingke Weng, Elizabeth TorresAbstract:Bioluminescence, or the production of light by living organisms via chemical reaction, is widespread across Metazoa. Culturing bioluminescent organisms from diverse taxonomic groups is important for determining the biosynthetic pathways of bioluminescent substrates, which may lead to new tools for biotechnology and biomedicine. Although some previously cultured luminescent groups represent independent origins of bioluminescence, cnidarians, ctenophores, and brittle stars use luminescent substrates (Luciferins) obtained from their diets, and therefore are not informative for determination of the biosynthethic pathways of the Luciferins. Terrestrial fireflies do synthesize their own Luciferin, but the biosynthetic pathway for firefly Luciferin remains unclear. An additional independent origin of endogenous bioluminescence is found within ostracods from the family Cypridinidae, which use their luminescence for defense and, in Caribbean species, for courtship displays. Here, we report the first complete life cycle of a luminous ostracod (Vargula tsujii, the California Sea Firefly) in the laboratory. We also describe the embryonic development of Vargula tsujii and discuss the size classes of instar development. We find embryogenesis in V. tsujii ranges between 25-38 days, and this species appears to have five instar stages, consistent with ontogeny in other cypridinid lineages. We estimate a complete life cycle at 3-4 months. We present the first complete mitochondrial genome for Vargula tsujii. Finally, we find no evidence of significant population genetic structure or cryptic species throughout the southern California range of V. tsujii. Bringing a luminous ostracod into laboratory culture sets the stage for many potential avenues of study, including the biosynthetic pathway of cypridinid Luciferin and genomic manipulation of an autogenic bioluminescent system.
Satoshi Inouye - One of the best experts on this subject based on the ideXlab platform.
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identification of 3 enol sulfate of cypridina Luciferin cypridina luciferyl sulfate in the sea firefly cypridina vargula hilgendorfii
Tetrahedron, 2014Co-Authors: Mitsuhiro Nakamura, Tomoko Suzuki, Norihiro Ishizaka, Junichi Sato, Satoshi InouyeAbstract:Abstract Cypridina Luciferin from the luminous ostracod Cypridina (Vargula) hilgendorfii has an imidazopyrazinone core structure (3,7-dihydroimidazopyrazin-3-one), which is identical to that of coelenterazine. Cypridina luciferyl sulfate (3-enol sulfate of Cypridina Luciferin) was isolated for the first time and the chemical structure was identified by LC/ESI–TOF–MS analysis. Furthermore, Cypridina luciferyl sulfate was chemically synthesized, and its absorption and MS/MS spectra were in agreement with that of Cypridina luciferyl sulfate isolated. Using the crude extracts of Cypridina specimens, Cypridina luciferyl sulfate could be converted to Cypridina Luciferin in the presence of adenosine 3′,5′-diphosphate (PAP), and Cypridina Luciferin was converted to Cypridina luciferyl sulfate in the presence of 3′-phosphoadenosine 5′-phosphosulfate (PAPS). These results suggested that a sulfotransferase catalyzes the reversible sulfation of Cypridina Luciferin in Cypridina hilgendorfii. In aqueous solution, Cypridina luciferyl sulfate was more stable than Cypridina Luciferin and might be a storage form of Cypridina Luciferin.
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Biosynthesis of Firefly Luciferin in Adult Lantern: Decarboxylation of ʟ-Cysteine is a Key Step for Benzothiazole Ring Formation in Firefly Luciferin Synthesis
PloS one, 2013Co-Authors: Yuichi Oba, Makoto Ojika, Naoki Yoshida, Shusei Kanie, Satoshi InouyeAbstract:Background Bioluminescence in fireflies and click beetles is produced by a luciferase-Luciferin reaction. The luminescence property and protein structure of firefly luciferase have been investigated, and its cDNA has been used for various assay systems. The chemical structure of firefly Luciferin was identified as the ᴅ-form in 1963 and studies on the biosynthesis of firefly Luciferin began early in the 1970’s. Incorporation experiments using 14C-labeled compounds were performed, and cysteine and benzoquinone/hydroquinone were proposed to be biosynthetic component for firefly Luciferin. However, there have been no clear conclusions regarding the biosynthetic components of firefly Luciferin over 30 years. Methodology/Principal Findings Incorporation studies were performed by injecting stable isotope-labeled compounds, including ʟ-[U-13C3]-cysteine, ʟ-[1-13C]-cysteine, ʟ-[3-13C]-cysteine, 1,4-[D6]-hydroquinone, and p-[2,3,5,6-D]-benzoquinone, into the adult lantern of the living Japanese firefly Luciola lateralis. After extracting firefly Luciferin from the lantern, the incorporation of stable isotope-labeled compounds into firefly Luciferin was identified by LC/ESI-TOF-MS. The positions of the stable isotope atoms in firefly Luciferin were determined by the mass fragmentation of firefly Luciferin. Conclusions We demonstrated for the first time that ᴅ- and ʟ-firefly Luciferins are biosynthesized in the lantern of the adult firefly from two ʟ-cysteine molecules with p-benzoquinone/1,4-hydroquinone, accompanied by the decarboxylation of ʟ-cysteine.
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Determination of the Luciferin contents in luminous and non-luminous beetles.
Bioscience biotechnology and biochemistry, 2008Co-Authors: Yuichi Oba, Makoto Ojika, Takeru Shintani, Takanori Nakamura, Satoshi InouyeAbstract:The contents of firefly Luciferin in luminous and non-luminous beetles were determined by the methods of HPLC with fluorescence detection and the luminescence reaction of Luciferin and firefly luciferase. Luminous cantharoids and elaterids contained various amounts of Luciferin in the range of pmol to hundreds of nmol, but no Luciferin was detected in the non-luminous cantharoids and elaterids.
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Stereoselective Incorporation of Isoleucine into Cypridina Luciferin in Cypridina hilgendorfii (Vargula hilgendorfii)
Bioscience biotechnology and biochemistry, 2006Co-Authors: Shin-ichi Kato, Yuichi Oba, Makoto Ojika, Satoshi InouyeAbstract:The emission of light in the marine ostracod Cypridina hilgendorfii (presently Vargula hilgendorfii) is produced by the Cypridina Luciferin-luciferase reaction in the presence of molecular oxygen. Cypridina Luciferin has an asymmetric carbon derived from isoleucine, and the absolute configuration is identical to the C-3 position in L-isoleucine or D-alloisoleucine. To determine the stereoselective incorporation of the isoleucine isomers (L-isoleucine, D-isoleucine, L-alloisoleucine, and D-alloisoleucine), we synthesized four 2H-labeled isoleucine isomers and examined their incorporation into Cypridina Luciferin by feeding experiments. Judging by these results, L-isoleucine is predominantly incorporated into Cypridina Luciferin. This suggests that the isoleucine unit of Cypridina Luciferin is derived from L-isoleucine, but not from D-alloisoleucine.
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Fusions to imidazopyrazinone-type luciferases and aequorin as reporters.
Methods in Enzymology, 2004Co-Authors: Satoshi InouyeAbstract:Publisher Summary This chapter discusses the fusion of genes to imidazopyrazinone-type luciferases and aequorin as reporters. In bioluminescent marine organisms, an imidazopyrazinone (3,7-dihydroimidazopyrazin-3-one) compound is used for the luciferase reaction. Two imidazopyrazinone-type Luciferins, coelenterazine Luciferin and Cypridina Luciferin, have been isolated. These two Luciferins are similar with a central imidazolepyrazine nucleus in their structures. Coelenterazine is also known as “Watasenia preLuciferin,” “Oplophorus Luciferin,” and “Renilla Luciferin.” Among coelenterazine-type luciferases, Renilla luciferase and Oplophorus luciferase have been isolated and their biochemical properties characterized. Coelenterazine serves as the chromogenic compound of photoproteins—such as aequorin, phialidin, halistaurin, and obelin. In the field of molecular biology, several reporter genes are used for monitoring the gene expression in whole organisms, as well as in single cells. Luciferases, including firefly luciferase and photoproteins, are useful reporter molecules with various advantages for the bioluminescent system: they are sensitive, rapid, and harmless. Moreover, the detection of photon produced by luciferase permits the real-time imaging of gene expression and of the dynamic changes of target protein in living cells. The luminescence reaction of imidazopyrazinone-type luciferases is simpler in required components than that of firefly luciferase, because the imidazopyrazinone-type luciferase systems do not need any cofactors such as ATP and Mg 2+ .
Jennifer A. Prescher - One of the best experts on this subject based on the ideXlab platform.
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Multicomponent Bioluminescence Imaging with a π-Extended Luciferin
Journal of the American Chemical Society, 2020Co-Authors: Zi Yao, Brendan S. Zhang, Rachel C Steinhardt, Jeremy H. Mills, Jennifer A. PrescherAbstract:Bioluminescence imaging with luciferase-Luciferin pairs is commonly used for monitoring biological processes in cells and whole organisms. Traditional bioluminescent probes are limited in scope, though, as they cannot be easily distinguished in biological environments, precluding efforts to visualize multicellular processes. Additionally, many luciferase-Luciferin pairs emit light that is poorly tissue penetrant, hindering efforts to visualize targets in deep tissues. To address these issues, we synthesized a set of π-extended Luciferins that were predicted to be red-shifted luminophores. The scaffolds were designed to be rotationally labile such that they produced light only when paired with luciferases capable of enforcing planarity. A Luciferin comprising an intramolecular "lock" was identified as a viable light-emitting probe. Native luciferases were unable to efficiently process the analog, but a complementary luciferase was identified via Rosetta-guided enzyme design. The unique enzyme-substrate pair is red-shifted compared to well-known bioluminescent tools. The probe set is also orthogonal to other luciferase-Luciferin probes and can be used for multicomponent imaging. Four substrate-resolved luciferases were imaged in a single session. Collectively, this work provides the first example of Rosetta-guided design in engineering bioluminescent tools and expands the scope of orthogonal imaging probes.
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Pyridone Luciferins and Mutant Luciferases for Bioluminescence Imaging
ChemBioChem, 2018Co-Authors: Brendan S. Zhang, Krysten A. Jones, David C. Mccutcheon, Jennifer A. PrescherAbstract:: New applications for bioluminescence imaging require an expanded set of luciferase enzymes and Luciferin substrates. Here, we report two novel Luciferins for use in vitro and in cells. These molecules comprise regioisomeric pyridone cores that can be accessed from a common synthetic route. The analogues exhibited unique emission spectra with firefly luciferase, although photon intensities remained weak. Enhanced light outputs were achieved by using mutant luciferase enzymes. One of the Luciferin-luciferase pairs produced light on par with native probes in live cells. The pyridone analogues and complementary luciferases add to a growing set of designer probes for bioluminescence imaging.
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orthogonal luciferase Luciferin pairs for bioluminescence imaging
Journal of the American Chemical Society, 2017Co-Authors: Krysten A. Jones, David C. Mccutcheon, William B Porterfield, Colin M Rathbun, Miranda A Paley, Jennifer A. PrescherAbstract:Bioluminescence imaging with luciferase–Luciferin pairs is widely used in biomedical research. Several luciferases have been identified in nature, and many have been adapted for tracking cells in whole animals. Unfortunately, the optimal luciferases for imaging in vivo utilize the same substrate and therefore cannot easily differentiate multiple cell types in a single subject. To develop a broader set of distinguishable probes, we crafted custom Luciferins that can be selectively processed by engineered luciferases. Libraries of mutant enzymes were iteratively screened with sterically modified Luciferins, and orthogonal enzyme–substrate “hits” were identified. These tools produced light when complementary enzyme–substrate partners interacted both in vitro and in cultured cell models. Based on their selectivity, these designer pairs will bolster multicomponent imaging and enable the direct interrogation of cell networks not currently possible with existing tools. Our screening platform is also general and ...
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Orthogonal Luciferase–Luciferin Pairs for Bioluminescence Imaging
Journal of the American Chemical Society, 2017Co-Authors: Krysten A. Jones, David C. Mccutcheon, William B Porterfield, Colin M Rathbun, Miranda A Paley, Jennifer A. PrescherAbstract:Bioluminescence imaging with luciferase–Luciferin pairs is widely used in biomedical research. Several luciferases have been identified in nature, and many have been adapted for tracking cells in whole animals. Unfortunately, the optimal luciferases for imaging in vivo utilize the same substrate and therefore cannot easily differentiate multiple cell types in a single subject. To develop a broader set of distinguishable probes, we crafted custom Luciferins that can be selectively processed by engineered luciferases. Libraries of mutant enzymes were iteratively screened with sterically modified Luciferins, and orthogonal enzyme–substrate “hits” were identified. These tools produced light when complementary enzyme–substrate partners interacted both in vitro and in cultured cell models. Based on their selectivity, these designer pairs will bolster multicomponent imaging and enable the direct interrogation of cell networks not currently possible with existing tools. Our screening platform is also general and ...
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Design and Synthesis of an Alkynyl Luciferin Analogue for Bioluminescence Imaging
Chemistry (Weinheim an der Bergstrasse Germany), 2016Co-Authors: Rachel C Steinhardt, David C. Mccutcheon, Colin M Rathbun, Miranda A Paley, Jessica M. O'neill, Jennifer A. PrescherAbstract:Herein, the synthesis and characterization of an alkyne-modified Luciferin is reported. This bioluminescent probe was accessed using C-H activation methodology and was found to be stable in solution and capable of light production with firefly luciferase. The Luciferin analogue was also cell permeant and emitted more redshifted light than d-Luciferin, the native luciferase substrate. Based on these features, the alkynyl Luciferin will be useful for a variety of imaging applications.
