Determination of the Binding Parameters between Proteins and Luminol by Chemiluminescence Using Flow Injection Technique

Jie Guo,Zhenghua Song,Donghua Chen

doi:10.1155/2013/391053

Abstract

The interaction behavior of bovine serum albumin (BSA), lysozyme (LYS), myoglobin (MB), and catalase (CAT) with luminol, respectively, was first studied by chemiluminescence (CL) using flow injection (FI) technique based on the fact that the studied proteins can enhance the CL intensity of luminol. A FI-CL model of protein-luminol interaction, lg[(I0−I)/I]=1/nlg[P]+1/nlgKa+2lgn, was constructed, and the interaction parameters of BSA, LYS, MB, and CAT with luminol were determined accordingly. The binding constants Ka are in the descending order of CAT > MB > LYS > BSA at the level of 105 to 107 L mol−1, and the number of binding sites n of luminol to BSA or LYS is around 2 and to MB or CAT is around 1. The results of thermodynamic parameters (ΔH, ΔS, and ΔG) showed that the binding processes of luminol to the four proteins are spontaneous mainly through the hydrophobic force.

Highlights

Proteins possess many biological functions including binding, catalysis, operating as molecular switches, and serving as structural components of cells and organisms [1, 2] and play an important role in the transportation and deposition of various endogenous and exogenous substances [3, 4]
The interaction of protein with small molecules has become a hot spot in the fields of chemistry, biology, and medicine [5,6,7]
All reagents used were of analytical reagent grade unless otherwise specified and doubly deionized water was purified in a Milli-Q system (Millipore, Bedford, MA, USA) for the preparation of solutions in the whole procedure

Summary

Introduction

Proteins possess many biological functions including binding, catalysis, operating as molecular switches, and serving as structural components of cells and organisms [1, 2] and play an important role in the transportation and deposition of various endogenous and exogenous substances [3, 4]. Luminol (5-amino-2, 3-dihydro-1, 4-phthalazinedione) is known to produce CL with the characteristic wavelength of 425 nm under alkaline condition [20, 21], making it widely used around the world as a blood enhancement technique in forensic science. This luminescent compound is used clinically in the treatment of alopecia [22], the promotion of blood clotting [23], and wound healing [24]. The CL of luminol has been employed to study its interaction behavior with serum albumins [12, 29]

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