Few substitutions affect the bioluminescence spectra of Phrixotrix (Coleoptera: Phengodidae) luciferases: a site-directed mutagenesis survey

Phrixotrix (railroad worm) luciferases produce bioluminescence in the green and red regions of the spectrum, depending on the location of the lanterns, and are the only luciferases naturally producing red bioluminescence. Comparison of the luciferase sequences showed a set of substitutions that could be involved in bioluminescence colour determination: (a) unique substitutions in the red luciferase replacing otherwise invariant residues; (b) conserved basic residues in the green-yellow emitting luciferases; and (c) an additional R353 residue in red-emitting luciferase (Viviani et al., 1999). To investigate whether these sites have a functional role in bioluminescence colour determination, we performed a site-directed mutagenesis. Natural substitutions in the region 220-344 and residues in the putative luciferin-binding site were also investigated. With the exception of the previously identified substitution of R215 and T226 (Viviani et al., 2002), which display dramatic red-shift effects on the spectrum of green-yellow-emitting luciferases, only a few substitutions had a moderate effect on the spectrum of the green-emitting luciferase. In contrast, no single substitution affected the spectrum of the red-emitting luciferase. The results suggest that the identity of the active site residues is not so critical for determining red bioluminescence in PxRE luciferase. Rather, the conformation assumed during the emitting step could be critical to set up proper interactions with excited oxyluciferin. Copyright ©2007 John Wiley & Sons, Ltd.

The influence of the loop between residues 223-235 in beetle luciferase bioluminescence spectra: A solvent gate for the active site of pH-sensitive luciferases

Beetle luciferases emit a wide range of bioluminescence colors, ranging from green to red. Firefly luciferases can shift the spectrum to red in response to pH and temperature changes, whereas click beetle and railroadworm luciferases do not. Despite many studies on firefly luciferases, the origin of pH-sensitivity is far from being understood. Through comparative site-directed mutagenesis and modeling studies, using the pH-sensitive luciferases (Macrolampis and Cratomorphus distinctus fireflies) and the pH-insensitive luciferases (Pyrearinus termitilluminans, Phrixotrix viviani and Phrixotrix hirtus) cloned by our group, here we show that substitutions dramatically affecting bioluminescence colors in both groups of luciferases are clustered in the loop between residues 223-235 (Photinus pyralis sequence). The substitutions at positions 227, 228 and 229 (P. pyralis sequence) cause dramatic redshift and temporal shift in both groups of luciferases, indicating their involvement in labile interactions. Modeling studies showed that the residues Y227 and N229 are buried in the protein core, fixing the loop to other structural elements participating at the bottom of the luciferin binding site. Changes in pH and temperature (in firefly luciferases), as well as point mutations in this loop, may disrupt the interactions of these structural elements exposing the active site and modulating bioluminescence colors. © 2007 The Authors.

The structural origin and biological function of pH-sensitivity in firefly luciferases

Firefly luciferases are called pH-sensitive because their bioluminescence spectra display a typical red-shift at acidic pH, higher temperatures, and in the presence of heavy metal cations, whereas other beetle luciferases (click beetles and railroadworms) do not, and for this reason they are called pH-insensitive. Despite many studies on firefly luciferases, the origin of pH-sensitivity is far from being understood. This subject is revised in view of recent results. Some substitutions of amino-acid residues influencing pH-sensitivity in firefly luciferases have been identified. Sequence comparison, site-directed mutagenesis and modeling studies have shown a set of residues differing between pH-sensitive and pH-insensitive luciferases which affect bioluminescence colors. Some substitutions dramatically affecting bioluminescence colors in both groups of luciferases are clustered in the loop between residues 223-235 (Photinus pyralis sequence). A network of hydrogen bonds and salt bridges involving the residues N229-S284-E311-R337 was found to be important for affecting bioluminescence colors. It is suggested that these structural elements may affect the benzothiazolyl side of the luciferin-binding site affecting bioluminescence colors. Experimental evidence suggest that the residual red light emission in pH-sensitive luciferases could be a vestige that may have biological importance in some firefly species. Furthermore, the potential utility of pH-sensitivity for intracellular biosensing applications is considered. © The Royal Society of Chemistry and Owner Societies.

Pyrearinus termitilluminans larval click beetle luciferase: Active site properties, structure and function relationships and comparison with other beetle luciferases

Several beetle luciferases have been cloned and sequenced. However, most studies on structure and function relationships and bioanalytical applications were done with firefly luciferases, which are pH sensitive. Several years ago we cloned Pyrearinus termitilluminans larval click beetle luciferase, which displays the most blue-shifted bioluminescence among beetle luciferases and is pH insensitive. This enzyme was expressed in E. coli, purified, and its properties investigated. This luciferase shows slower luminescence kinetics, KM values comparable to other beetle luciferases and high catalytic constant. Fluorescence studies with 8-anilino-1-naphtalene-sulfonic acid (1,8-ANS) and modeling studies suggest that the luciferin binding site of this luciferase is very hydrophobic, supporting the solvent and orientation polarizability effects as determining mechanisms for bioluminescence colors. Although pH insensitive in the range between pH 6-8, at pH 10 this luciferase displays a remarkable red-shift and broadening of the bioluminescence spectrum. Modeling studies suggest that the residue C312 may play an important role in bioluminescence color modulation. Compared to other beetle luciferases, Pyrearinus termitilluminans luciferase also displays higher thermostability and sustained luminescence in a bacterial cell environment, which makes this luciferase particularly suitable for in vivo cell analysis and bioimaging. © The Royal Society of Chemistry and Owner Societies 2009.

Aspectos evolutivos da bioluminescência de elateroidea (coleoptera) do Brasil

Pós-graduação em Ciências Biológicas (Biologia Celular e Molecular) - IBRC

Determination of ages and fossil Coleoptera fauna from eight sites in the London region

These data sets report the fossil beetle assemblages identified from the Mesolithic to Late Bronze Age at eight sites in the London region. All but one of the study sites are within 2 km of the modern course of the Thames. The sites produced 128 faunal assemblages that yielded 218 identified species in 41 families of Coleoptera (beetles).  Beetle faunas of Mesolithic age indicate extensive wetlands near the Thames, bordered by rich deciduous woodlands. The proportion of woodland species declined in the Neolithic, apparently because of the expansion of wetlands, rather than because of human activities. The Early Bronze Age faunas contained a greater proportion of coniferous woodland and aquatic (standing water) species. An increase in the dung beetle fauna indicates the presence of sheep, cattle and horses, and various beetles associated with crop lands demonstrate the local rise of agriculture, albeit several centuries after the beginnings of farming in other regions of Britain. Late Bronze Age faunas show the continued development of agriculture and animal husbandry along the lower Thames. About 33% of the total identified beetle fauna from the London area sites have limited modern distributions or are extinct in the U.K. Some of these species are associated with the dead wood found in primeval forests; others are wetland species whose habitat has been severely reduced in recent centuries. The third group is stream-dwelling beetles that require clean, clear waters and river bottoms.

Monografía de los elatéridos de Chile

Mode of access: Internet.

Identificación y fluctuación poblacional de insectos asociados al cultivo de la piña (Ananas comosus L. Merril) Ticuantepe, Nicaragua.

Con el objetivo de contribuir al conocimiento científico de insectos asociadas al cultivo de la piña (Ananas comosus L. Merril), se realizó este estudio en el período de marzo a septiembre del 2014 en la zona piñera del Municipio de Ticuantepe, teniendo como objetivos específicos la identificación, abundancia, riqueza, índice de biodiversidad, fluctuación poblacional y el hábito de las principales familias de insectos asociados al cultivo de la piña. El estudio se desarrolló en seis fincas piñeras donde se ubicaron tres trampas de caída libre (Pitfall traps) para la captura de insectos rastreros, tres trampas de galones de plástico con melaza para la captura de insectos voladores y en cada sitio se colectó manualmente insectos del follaje frutos y suelo. En esta investigación se encontró que las principales familias asociadas al cultivo son; Formicidae, Scarabaeidae, Pseudococcidae, Elateridae, Noctuidae, Nymphalidae Cerambicidae y Lycaenidae. La diversidad insectil se estimó utilizando el índice de diversidad Shannon-Weaver, la familia con mayor índice de diversidad fue Formicidae, En cuanto a la riqueza de insectos se encontraron cincuenta y un familias y ochenta y cuatro géneros, siendo los más numerosos pertenecientes a las familias; Formicidae, Scarabaidae, Tenebrionidae y Pentatomidae. En lo que respecta a la fluctuación poblacional, las familias de insectos que presentaron mayor incidencia fueron: Formicidae, Scarabaidae y Pseudococcidae, las fluctuaciones más altas de insectos ocurrieron en los meses de abril, mayo y julio, entre los principales hábitos alimenticios de las especies insectiles asociados al cultivo de la piña se encontraron, defoliadores, plagas del suelo, succionadores de floemas, barrenadores de corteza vegetal, polinizadoras, depredadores, parasitoides y degradadores de materia orgánica.