DNA binding, protein interaction and nuclease activity of a new Cu(II)-N-substituted sulfonamide complex

Tamara Liana Topală,a Alejandro Pascual-Álvarez,b Alfonso Castiñeiras,c Andreea Bodoki,a Luminiţa Oprean,a Radu Oprean,d Gloria Alzuet-Piñab

a) Department of General and Inorganic Chemistry, Faculty of Pharmacy, UMF “Iuliu Haţieganu” Cluj-Napoca, Romania; b) Department of Inorganic Chemistry, Faculty of Pharmacy, Universitat de Valencia, Spain;

c) Department of Inorganic Chemistry, University of Santiago de Compostela, Santiago de Compostela, Spain; d) Department of Analytical Chemistry and Instrumental Analysis, Faculty of Pharmacy, UMF “Iuliu Haţieganu” Cluj-Napoca, Romania

topala.liana@umfcluj.ro

AcknowledgmentsT.L.T., A.B., L.O. and R.O. acknowledge the financial support offered by research Project POSDRU 107/1.5/S/78702. A.P.A and G.A.P. acknowledge financial support from the Spanish Comisión Interministerial de Ciencia y Tecnología (CTQ2007–63690/BQU).

References[1] C.–L. Liu, M. Wang, T.–L. Zhang, H.–Z. Sun, Coord. Chem. Rev. 248 (2004) 147–168; [2] P. Sathyadevi, P. Krishnamoorthy, E. Jayanthi, R.R. Butorac, A.H. Cowley, N. Dharmaraj. Inorganica Chimica Acta 384 (2012) 83-96. [3] D.S. Sigman, D.R. Graham, V. Daurora, A.M. Stern, J. Biol. Chem. 254 (1979) 12269–12272; [4] M. Oivanen, S. Kuusela, H. Lonnberg, Chem. Rev. 98 (1998) 961–990.

Interaction with biomacromolecules

ConclusionsThe newly synthesized Cu(II) complex shows a relatively high affinity towards biomacromolecules, as shown by the fluorescence studies, and a significant nuclease activity in the presence of ascorbic acid as well as upon irradiation with ultra-violet light.

DNA interaction

DNA thermal denaturation

Interaction with the EtBr-DNA system

[Cu(L)Cl]n crystal structure

DNA cleavage

Introduction

Agarose gel electrophoresis of pUC18 plasmid treated with the complex; buffer: cacodylate 0.1 M, pH = 6; photoactivation: λ = 300-400 nm.

Photocleavage

SC

DN

A(n

on-ir

radi

ated

)

λDN

A/E

coR

I +H

indI

II

The ample potential applications of DNA cleavage agents in molecular biology, biotechnology, and medicine [1] sparked interest in designing new molecules with this property. Studying the binding of molecules to albumins is a very pertinent research area, taking into consideration their crucial roles in drug pharmacodynamics and pharmacokinetics [2].Since the discovery of the first chemical nuclease [3], increasing attention was focused on metal complexes as nuclease mimics because of their advantages in electronic and structural diversities [4].The objectives of the present work were to synthesize and characterize a new copper(II) complex with N-(pyridin-2-ylmethyl)quinoline-8-sulfonamide (HL) and explore its ability to bind to biomolecules (DNA and bovine serum albumin, BSA), as well as its nuclease activity.

Emission spectrum of EtBr bound to DNA in the absence and presence of increasing

concentrations of [Cu(L)Cl]n. The arrow shows the effect of the complex on the emission intensity of

the DNA-EtBr aduct

The decrease in the emission intensity of the EtBr-DNA system indicates that the complex binds to DNA.

Bovine serum albumin interaction

[DNA] = 50 µM[EtBr] = 50 µM5 % DMF

21 %

[Cu(L)Cl]n :

[Cu(L)Cl]n / [BSA] : 40 %

Fluorescence quenching spectrum of BSA as a function of concentration of [Cu(L)Cl]n. The

arrow shows the effect of the complex on the emission intensity of BSA.

[BSA] = 1.3 µM5 % DMF

The decrease in the fluorescence emission intensity is due to changes in the environment of the tryptophan moieties in the flourophore caused by the binding of [Cu(L)Cl]n.

KSV = 2.6 x 105 M-1

Kb = 1.4 x 105 M-1

n = 0.95

τ = 0.151T5 = 0.887 slightly distorted square pyramidal geometry

N(18)-Cu(1)-N(11) 82.37(5) N(18)-Cu(1)-N(27) 88.81(5)N(11)-Cu(1)-N(27) 170.88(5)N(18)-Cu(1)-Cl(1) 161.82(4)N(11)-Cu(1)-Cl(1) 95.53(4)N(27)-Cu(1)-Cl(1) 93.58(4)N(18)-Cu(1)-O(11) 101.84(5)N(11)-Cu(1)-O(11) 93.61(5)N(27)-Cu(1)-O(11) 85.87(5)Cl(1)-Cu(1)-O(11) 96.31(3)

Selected bonds and angles

Cu(1)-N(18) 1.9817(13)Cu(1)-N(11) 2.0282(13)Cu(1)-N(27) 2.0494(13)Cu(1)-Cl(1) 2.2571(4)Cu(1)-O(11) 2.3431(11)Cu(1)-O(11)#1 2.8882(11)

Cu(1)-Cu(1)#1 5.0738(3)

20 25 30 40 50

SC

DN

A

CuCl2

20 25 30 40 50

[Cu(L)Cl]n

µMSupercoiled DNA

Open circular DNA

The difference between the nuclease activity shown by the complex and that of the Cu(II) salt is observed at lower concentrations (20-25 µM).

Nuclease activity in the presence of ascorbic acid

Agarose gel electrophoresis of pUC18 plasmid treated with the complex; buffer: cacodylate 0.1 M, pH = 6; ascorbic acid: 1x.

SC

DN

A

SC

DN

A +

as

corb

ic a

cid

λDN

A/E

coR

I +H

indI

II

10 15 20 25 30 10 15 20 25 30

CuCl2 [Cu(L)Cl]n

µM

There is a significant difference between the nuclease activity shown by the complex and that of the Cu(II) salt in the presence of ascorbic acid.

[Cu(L)Cl]n

[DNA] = 100 µM[Complex] = 50 µM5 % DMF

ΔTm = 7 °C

Thermal denaturation of CT-DNA in the absence and presence of [Cu(L)Cl]n

The Cu(II) complex stabilizes the DNA structure, increasing its melting point.