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A fusion protein designed in order to combine the fluorescence emission of the Green Fluorescent Protein (GFP) with the adhesion ability of the class I hydrophobin Vmh2 was heterologously produced in the yeast Pichia pastoris. The Vmh2-GFP fusion protein has proven to be a smart and effective tool for the study of Vmh2 self-assembling.Since the two proteins were linked by the specific cutting site of the thrombin, the fusion protein was used as the active biological element in the realization of a thrombin biosensor. When the thrombin present in the target solution specifically hydrolyzed its cleavage sequence, a consequent decrease in the fluorescence intensity of the sample could be observed. The Vmh2-GFP based assay allowed quantification of thrombin in solution with a detection limit of 2.27 aM. The specificity of the assay with respect to other proteases and proteins granted the measurement of thrombin …
Publication date: 
15 Jan 2017

Alessandra Piscitelli, Anna Pennacchio, Paola Cicatiello, Jane Politi, Luca De Stefano, Paola Giardina

Biblio References: 
Volume: 87 Pages: 816-822
Biosensors and Bioelectronics