A unique fluorescence response of hexaphenylsilole to methyl parathion hydrolase: a new signal generating system for the enzyme label

Abstract

Methyl parathion hydrolase (MPH), an enzyme that catalyses the turnover of methyl parathion (MP) to p-nitrophenol (pNP), can be utilized as an enzyme label. In this paper, a unique fluorescence response of 1,1-bis[4-(diethylaminomethyl)phenyl]-2,3,4,5-tetraphenylsilole (A₂HPS) to MPH whose gene was obtained from Pseudomonas sp. strain WBC-3 is described. In the absence of MP, A₂HPS could only give a small fluorescence response to the enzyme (I/I₀ = 1.1). The detection must be performed under low pH conditions, and the influence of BSA and hemoglobin (Hb) was high; upon addition of the enzyme's substrate, 1 × 10⁻⁵ μg mL⁻¹ or 2.85 × 10⁻¹³ M of the MPH could be reported by A₂HPS with a higher I/I₀ of 1.7. The detection limit was 10⁵ times more sensitive than that given by a spectrophotometric method. In addition, the assay could be performed at a near neutral pH which was more biocompatible, and little influence was observed from BSA and Hb. The light-on response to the MPH was due to the different quenching effect of the MP and pNP on A₂HPS and the improvement in the detection selectivity was due to combining the enzyme reaction with the detection. The findings of this work suggested that A₂HPS and MP could form a new reporter system for the MPH enzyme label.