Dataset Information

Mutagenesis and Structural Studies Reveal the Basis for the Activity and Stability Properties That Distinguish the Photinus Luciferases scintillans and pyralis.

ABSTRACT: The dazzling yellow-green light emission of the common North American firefly Photinus pyralis and other bioluminescent organisms has provided a wide variety of prominent research applications like reporter gene assays and in vivo imaging methods. While the P. pyralis enzyme has been extensively studied, only recently has a second Photinus luciferase been cloned from the species scintillans. Even though the enzymes share very high sequence identity (89.8%), the color of the light they emit, their specific activity and their stability to heat, pH, and chemical denaturation are quite different with the scintillans luciferase being generally more resistant. Through the construction and evaluation of the properties of chimeric domain swapped, single point, and various combined variants, we have determined that only six amino acid changes are necessary to confer all of the properties of the scintillans enzyme to wild-type P. pyralis luciferase. Altered stability properties were attributed to four of the amino acid changes (T214N/S276T/H332N/E354N), and single mutations each predominantly changed emission color (Y255F) and specific activity (A222C). Results of a crystallographic study of the P. pyralis enzyme containing the six changes (Pps6) provide some insight into the structural basis for some of the documented property differences.

SUBMITTER: Branchini BR

PROVIDER: S-EPMC6810810 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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Mutagenesis and Structural Studies Reveal the Basis for the Activity and Stability Properties That Distinguish the Photinus Luciferases scintillans and pyralis.

Branchini Bruce R BR Fontaine Danielle M DM Southworth Tara L TL Huta Brian P BP Racela Allison A Patel Ketan D KD Gulick Andrew M AM

Biochemistry 20191010 42

The dazzling yellow-green light emission of the common North American firefly Photinus pyralis and other bioluminescent organisms has provided a wide variety of prominent research applications like reporter gene assays and in vivo imaging methods. While the P. pyralis enzyme has been extensively studied, only recently has a second Photinus luciferase been cloned from the species scintillans. Even though the enzymes share very high sequence identity (89.8%), the color ...[more]

PMID: 31560532

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Project description:BACKGROUND:Genes underlying signal production and reception are expected to evolve to maximize signal detection in specific environments. Fireflies vary in their light signal color both within and between species, and thus provide an excellent system in which to study signal production and reception in the context of signaling environments. Differences in signal color have been hypothesized to be due to variation in the sequence of luciferase, the enzyme that catalyzes the light reaction. Similarly, differences in visual sensitivity, which are expected to match signal color, have been hypothesized to be due to variation in the sequence of opsins, the protein component of visual pigments. Here we investigated (1) whether sequence variation in luciferase correlates with variation in signal color and (2) whether sequence variation in opsins correlates with inferred matching visual sensitivity across populations of a widespread North American firefly species, Photinus pyralis. We further tested (3) whether selection has acted on these loci by examining their population-level differentiation relative to the distribution of differentiation derived from a genome-wide sample of loci generated by double-digest RADseq. RESULTS:We found virtually no coding variation in luciferase or opsins. However, there was extreme divergence in non-coding variation in luciferase across populations relative to a panel of random genomic loci. CONCLUSIONS:The absence of protein variation at both loci challenges the paradigm that variation in signal color and visual sensitivity in fireflies is exclusively due to coding variation in luciferase and opsin genes. Instead, flash color variation within species must involve other mechanisms, such as abdominal pigmentation or regulation of light organ physiology. Evidence for selection at non-coding variation in luciferase suggests that selection is targeting luciferase regulation and may favor differ expression levels across populations.

Dataset Information

Mutagenesis and Structural Studies Reveal the Basis for the Activity and Stability Properties That Distinguish the Photinus Luciferases scintillans and pyralis.

Publications

Mutagenesis and Structural Studies Reveal the Basis for the Activity and Stability Properties That Distinguish the <i>Photinus</i> Luciferases <i>scintillans</i> and <i>pyralis</i>.

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