Limits...
Binding of a fluorescence reporter and a ligand to an odorant-binding protein of the yellow fever mosquito, Aedes aegypti.

Leal GM, Leal WS - F1000Res (2014)

Bottom Line: Binding assays using the fluorescence reporter N-phenyl-1-naphtylamine (NPN) were inconclusive.However, titration of NPN fluorescence emission in AaegOBP1 solution with MOP led to unexpected and intriguing results.Quenching was observed in the initial phase of titration, but addition of higher doses of MOP led to a stepwise increase in fluorescence emission coupled with a blue shift, which can be explained at least in part by formation of MOP micelles to house stray NPN molecules.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, University of California, Davis, Davis, CA, 95616, USA ; Davis Senior High School, Davis, CA, 95616, USA.

ABSTRACT
Odorant-binding proteins (OBPs), also named pheromone-binding proteins when the odorant is a pheromone, are essential for insect olfaction. They solubilize odorants that reach the port of entry of the olfactory system, the pore tubules in antennae and other olfactory appendages. Then, OBPs transport these hydrophobic compounds through an aqueous sensillar lymph to receptors embedded on dendritic membranes of olfactory receptor neurons. Structures of OBPs from mosquito species have shed new light on the mechanism of transport, although there is considerable debate on how they deliver odorant to receptors. An OBP from the southern house mosquito, Culex quinquefasciatus, binds the hydrophobic moiety of a mosquito oviposition pheromone (MOP) on the edge of its binding cavity. Likewise, it has been demonstrated that the orthologous protein from the malaria mosquito binds the insect repellent DEET on a similar edge of its binding pocket. A high school research project was aimed at testing whether the orthologous protein from the yellow fever mosquito, AaegOBP1, binds DEET and other insect repellents, and MOP was used as a positive control. Binding assays using the fluorescence reporter N-phenyl-1-naphtylamine (NPN) were inconclusive. However, titration of NPN fluorescence emission in AaegOBP1 solution with MOP led to unexpected and intriguing results. Quenching was observed in the initial phase of titration, but addition of higher doses of MOP led to a stepwise increase in fluorescence emission coupled with a blue shift, which can be explained at least in part by formation of MOP micelles to house stray NPN molecules.

No MeSH data available.


Related in: MedlinePlus

Titration of NPN fluorescence emission with MOP.(A) NPN (2.5 µM) was added to a solution of AaegOBP1 (0.35 µM) in ammonium acetate buffer, pH 7. NPN (2.5 µM) was added to ammonium acetate buffer, (B) pH 7 or (C) pH 5.5.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4309172&req=5

f5: Titration of NPN fluorescence emission with MOP.(A) NPN (2.5 µM) was added to a solution of AaegOBP1 (0.35 µM) in ammonium acetate buffer, pH 7. NPN (2.5 µM) was added to ammonium acetate buffer, (B) pH 7 or (C) pH 5.5.

Mentions: Fluorescence reporter wasN-phenyl-1-naphthylamine (NPN). Insect repellents used were DEET, PMD, Picaridin and IR3535. Mosquito oviposition pheromone was used as a positive control. Please see ReadMe file for details regarding each file. Please see the associated article for methods. The raw data forFigure 5C has been added in this version.


Binding of a fluorescence reporter and a ligand to an odorant-binding protein of the yellow fever mosquito, Aedes aegypti.

Leal GM, Leal WS - F1000Res (2014)

Titration of NPN fluorescence emission with MOP.(A) NPN (2.5 µM) was added to a solution of AaegOBP1 (0.35 µM) in ammonium acetate buffer, pH 7. NPN (2.5 µM) was added to ammonium acetate buffer, (B) pH 7 or (C) pH 5.5.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4309172&req=5

f5: Titration of NPN fluorescence emission with MOP.(A) NPN (2.5 µM) was added to a solution of AaegOBP1 (0.35 µM) in ammonium acetate buffer, pH 7. NPN (2.5 µM) was added to ammonium acetate buffer, (B) pH 7 or (C) pH 5.5.
Mentions: Fluorescence reporter wasN-phenyl-1-naphthylamine (NPN). Insect repellents used were DEET, PMD, Picaridin and IR3535. Mosquito oviposition pheromone was used as a positive control. Please see ReadMe file for details regarding each file. Please see the associated article for methods. The raw data forFigure 5C has been added in this version.

Bottom Line: Binding assays using the fluorescence reporter N-phenyl-1-naphtylamine (NPN) were inconclusive.However, titration of NPN fluorescence emission in AaegOBP1 solution with MOP led to unexpected and intriguing results.Quenching was observed in the initial phase of titration, but addition of higher doses of MOP led to a stepwise increase in fluorescence emission coupled with a blue shift, which can be explained at least in part by formation of MOP micelles to house stray NPN molecules.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Biology, University of California, Davis, Davis, CA, 95616, USA ; Davis Senior High School, Davis, CA, 95616, USA.

ABSTRACT
Odorant-binding proteins (OBPs), also named pheromone-binding proteins when the odorant is a pheromone, are essential for insect olfaction. They solubilize odorants that reach the port of entry of the olfactory system, the pore tubules in antennae and other olfactory appendages. Then, OBPs transport these hydrophobic compounds through an aqueous sensillar lymph to receptors embedded on dendritic membranes of olfactory receptor neurons. Structures of OBPs from mosquito species have shed new light on the mechanism of transport, although there is considerable debate on how they deliver odorant to receptors. An OBP from the southern house mosquito, Culex quinquefasciatus, binds the hydrophobic moiety of a mosquito oviposition pheromone (MOP) on the edge of its binding cavity. Likewise, it has been demonstrated that the orthologous protein from the malaria mosquito binds the insect repellent DEET on a similar edge of its binding pocket. A high school research project was aimed at testing whether the orthologous protein from the yellow fever mosquito, AaegOBP1, binds DEET and other insect repellents, and MOP was used as a positive control. Binding assays using the fluorescence reporter N-phenyl-1-naphtylamine (NPN) were inconclusive. However, titration of NPN fluorescence emission in AaegOBP1 solution with MOP led to unexpected and intriguing results. Quenching was observed in the initial phase of titration, but addition of higher doses of MOP led to a stepwise increase in fluorescence emission coupled with a blue shift, which can be explained at least in part by formation of MOP micelles to house stray NPN molecules.

No MeSH data available.


Related in: MedlinePlus