Study on the interaction between 4-(1H-indol-3-yl)-2-(p-tolyl)quinazoline-3-oxide and human serum albumin

https://doi.org/10.1016/j.bmc.2020.115720Get rights and content

Abstract

An organic small-molecular drug, 4-(1H-indol-3-yl)-2-(p-tolyl)quinazoline-3-oxide 1a was synthesized. It was employed to investigate the binding interaction and mechanism with human serum albumin (HSA). The experimental results indicated that the fluorescence quenching of HSA by 1a is a static quenching process and formation 1a-HSA complex. The site competition experiments revealed that the combination of 1a on HSA are hydrophobic interactions in the IIA domain and hydrogen bonds in IIIA domain of HSA, and the hydrophobic interactions of 1a on HSA are stronger than that of hydrogen bonds. These results were also confirmed by molecular docking theoretic analysis and ANS-hydrophobic fluorescent probe experiment. Synchronous fluorescence experiments showed that the polarity of HSA microenvironment was increase in the interaction process of 1a with HSA. The results of binding distance explored indicated that the combination distance between 1a and HSA is 3.63 nm, which is between 0.5R0 and 1.5R0, revealing the energy transfer between HSA and 1a is non-radiative. These results are very helpful for people to screen out high efficient indoloquinazoline drugs.

Introduction

The interactions between bio-macromolecules and organic small-molecular compounds have attracted increasing interest in recent years.1, 2, 3, 4 Among biomacromolecules, human serum albumin (HSA) is a kind of rich source of soluble protein in plasma and play an important role in the transportation and deposition of endogenous and exogenous substances such as fatty acids, amino acids and drugs.5 Thus, HSA is usually selected as model protein for studying the interaction between ligands (organic small-molecular drugs) and macromolecules for evaluation their drug effect.6, 7 HSA is a circulating protein composed of 585 amino acid residues in heart-shaped form. The crystal structure of HSA showed that it is composed of three structural domains: IA domain is composed of 1–195 amino acid residues, IIA domain is composed of 196–383 amino acid residues, and IIIA domain is composed of 384–585 amino acids. There are two binding sites, site I and site II in IIA and IIIA. There is a large hydrophobic cavity in the IIA domain,8 which is conducive to ligand binding sites. In 1976, Sudlow et al. found that warfarin could be combined with site I and site II.9 In 1995, Theodore Peters found that digitoxin could bind to HSA, but not at site I and site II. After experimental verification, the binding site was defined as site III.10 Therefore, the subsequent researchers conducted three sites competition experiments with drugs to determine the binding sites of ligand with HSA. The interaction between ligand and protein determines the bioavailability of the ligand to serum protein, and can reasonably evaluate biological and physicochemical properties of ligand. At the same time, it can also provide more information for improving the characteristic of biological macromolecules. The relationship between drug development and human health is very close. Therefore, in the follow-up experiments, HSA is directly used as the experimental material for the study of the structure–activity relationship of small molecule drug.

Quinazoline11, 12, 13 and indole14, 15, 16 derivatives are useful nitrogen-containing heterocyclic compounds, which are core structural motifs of many natural products and pharmaceuticals. Heterocyclic compounds bearing these two skeletons most likely possess interesting biological and physicochemical properties.17 For example, indoloquinazoline derivatives have been reported to be protein kinase CK2 inhibitors18 and poly(ADP-ribose) polymerase-1(PARP-1) inhibitors.19 4-(indole-3-yl)quinazolines have been reported to be potent epidermal growth factor receptor tyrosine kinase inhibitors.20 Recently, our group has been studied the structure–activity relationship of some quinazoline compounds, with the aim of screening out a quinazoline-based pharmaceutical molecular.21, 22, 23 More recently, a serial of 4-(indole-3-yl)quinazoline-3-oxides were synthesized via a cross-dehydrogenative coupling of quinazoline-3-oxides and indoles in our laboratory (Scheme 1).24 By primary experiments, 1-H-indol-3-yl-2-(p-tolyl)quinazoline-3-oxide 1a was found had an obvious activation effect on tyrosinase, and its semi-activated mass concentration (EC50) was 0.13 ± 0.1 mM. Considering the potential medicinal use of this organic compound and the important physiological functions of HSA, here the interactions and the mechanism btween 1a and HAS were explored.

Section snippets

Reagents and materials

HSA and digitoxin were purchased from Sigma-Aldrich, USA. DMSO, anhydrous ethanol, disodium hydrogen phosphate, sodium dihydrogen phosphate, etc. were purchased from Shanghai Sinopharm group. Ultrapure water was used throughout the experiment.

1-H-indol-3-yl-2-(p-tolyl)quinazoline-3-oxide 1a was synthesized via a cross-dehydrogenative coupling of quinazoline-3-oxide and indole according to our reported (scheme 1).24 The experimental procedures are as follow: 2-(p-tolyl)quinazoline-3-oxide

Fluorescence quenching analysis the interaction between HSA and compound 1a

As shown in Fig. 1A, the fluorescence emission of HSA has the maximum value at the wavelength of 333 nm (curve 1), and curve a is the fluorescence of compound 1a itself. As the concentration of 1a increased, the fluorescence absorption at 333 nm decreased significantly. As shown in Fig. 1B, when the concentration of 1a increased from 0 to 100 μM, the fluorescence intensity decreased by 49.2%, indicating that 1a can be bind with HSA and results fluorescence quenching.

In order to elucidate the

Conclusion

The interaction and mechanism of 4-(1H-indol-3-yl)-2-(p-tolyl)quinazoline-3-oxide 1a on HSA were investigated. Fluorescence quenching experiments showed that the quenching of HSA by 1a was static quenching, the combination of 1a and HSA resulted in the fluorescence quenching of HSA. Site competition experimental and molecular docking results show that the combination site of 1a on HSA is hydrophobic interaction in the IIA structure domain and is hydrogen bonds in the IIIA structure domain. The

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

Financial support from National Natural Science Foundation of China (no. 21762020 and 21362014), Jiangxi Provincial Department of Science and Technology (no. 20171BAB203006), and Key Laboratory of Functional Small Organic Molecule, Ministry of Education, Jiangxi Normal University (no. KLFS-KF-201623) is gratefully acknowledged.

References (45)

  • Y. Rao et al.

    Discovery of natural alkaloid bouchardatine as a novel inhibitor of adipogenesis/lipogenesis in 3T3-L1 adipocytes

    Bioorg Med Chem

    (2015)
  • M.K. Wei et al.

    Quinazolinone derivatives: Synthesis and comparison of inhibitory mechanisms on α-glucosidase

    Bioorg Med Chem

    (2017)
  • R. Wang et al.

    2-(4-Fluorophenyl)-quinazolin-4(3H)-one as a novel tyrosinase inhibitor: Synthesis, inhibitory activity, and mechanism

    Bioorg Med Chem

    (2016)
  • Y.Z. Zhang et al.

    Interaction of malachite green with bovine serum albumin: Determination of the binding mechanism and binding site by spectroscopic methods

    J Hazard Mater

    (2009)
  • Z.X. Chi et al.

    Toxic interaction mechanism between oxytetracycline and bovine hemoglobin

    J Hazard Mater

    (2010)
  • W.D. Horrocks et al.

    Measurement of distance between fluorescent amino acid residues and metal ion binding sites. Quantitation of energy transfer between tryptophan and terbium (III) or europium (III) in thermolysin

    Biochem Biophys Res Commun

    (1981)
  • X.C. Zhao et al.

    The interaction between Ag+ and bovine serum albumin: A spectroscopic investigation

    Sci Total Environ

    (2011)
  • O.K. Gasymov et al.

    ANS fluorescence: Potential to augment the identification of the external binding sites of proteins

    Biochim Biophys Acta

    (2007)
  • D. Matulis et al.

    1-Anilino-8-naphthalene sulfonate anion-protein binding depends primarily on ion pair formation

    Biophys J

    (1998)
  • N. Shahabadi et al.

    Multispectroscopic studies on the interaction of 2-tert-butylhydroquinone (TBHQ), a food additive, with bovine serum albumin

    Food Chem

    (2011)
  • J.R. Lakowicz et al.

    On the possibility of long-wavelength long-lifetime high-quantum-yield luminophores

    Anal Biochem

    (2001)
  • R.G. Gouveiaa et al.

    Synthesis, DNA and protein interactions and human topoisomerase inhibition of novel Spiro-acridine derivatives

    Bioorg Med Chem

    (2018)
  • Cited by (0)

    View full text