Tài liệu Báo cáo khoa học: Inhibition of cobalamin-dependent methionine synthase by substituted

Inhibition of cobalamin-dependent methionine synthase

by substituted benzo-fused heterocycles

Elizabeth C. Banks1

, Stephen W. Doughty2,*, Steven M. Toms1

, Richard T. Wheelhouse1

and Anna Nicolaou1

1 School of Pharmacy, University of Bradford, UK

2 School of Pharmacy, Centre for Biomolecular Sciences, University of Nottingham, UK

Methionine synthase (MetS) (5-methyltetrahydrofolate￾1homocysteine transmethylase) (EC.2.1.1.13) is one of

two established mammalian enzymes that utilize a bio￾logically active cobalamin derivative [methylcobalamin

(CH3-Cbl)] as a cofactor [1]. MetS catalyses the transfer

of the methyl group from 5-methyltetrahydrofolate to

homocysteine via the CH3-Cbl cofactor, with cycling

of cobalamin between the +1 [Cbl(I)] and +3 [Cbl(III)]

valency states (Fig. 1). Studies on the Escherichia coli

and Homo sapiens cobalamin-dependent MetS have

revealed that it is a large, conformationally flexible pro￾tein, consisting of four functional domains arranged

in a linear manner. Each one of these domains binds a

different substrate or cofactor. In detail, the N-terminal

module is the homocysteine (Hcy)-binding domain; the

second domain binds 5-methyltetrahydrofolate, the third

domain binds CH3-Cbl; and the fourth domain (C-term￾inal module) binds S-adenosyl-methionine (S-AdoMet),

an allosteric cofactor required for reductive reactivation

[2]. X-ray crystal structures of the cobalamin-, S-AdoMet￾and 5-methyltetrahydrofolate-binding sites have only

been reported for the bacterial enzyme [3–5].

The reaction products methionine and tetrahydro￾folate are further metabolized through the one-carbon

methionine transmethylation and folate cycles. MetS

is therefore intimately linked to important biochemical

Keywords

benzimidazole; benzothiadiazole; inhibition;

methionine synthase; molecular modelling

Correspondence

A. Nicolaou, School of Pharmacy, University

of Bradford, Richmond Road, Bradford BD7

1DP, UK

Fax: +44 1274 235600

Tel: +44 1274 234717

E-mail: [email protected]

*Present address

Faculty of Health and Biological Sciences,

School of Pharmacy, University of Notting￾ham Malaysia Campus, Jalan Broga, 43500

Semenyih, Selangor Darul Ehsan, Malaysia

(Received 14 July 2006, revised 7 November

2006, accepted 9 November 2006)

doi:10.1111/j.1742-4658.2006.05583.x

The cobalamin–dependent cytosolic enzyme, methionine synthase

(EC.2.1.1.13), catalyzes the remethylation of homocysteine to methionine

using 5-methyltetrahydrofolate as the methyl donor. The products of this

remethylation – methionine and tetrahydrofolate – participate in the active

methionine and folate pathways. Impaired methionine synthase activity has

been implicated in the pathogenesis of anaemias, cancer and neurological

disorders. Although the need for potent and specific inhibitors of methion￾ine synthase has been recognized, there is a lack of such agents. In this

study, we designed, synthesized and evaluated the inhibitory activity of a

series of substituted benzimidazoles and small benzothiadiazoles. Kinetic

analysis revealed that the benzimidazoles act as competitive inhibitors of

the rat liver methionine synthase, whilst the most active benzothiadiazole

(IC50 ¼ 80 lm) exhibited characteristics of uncompetitive inhibition. A

model of the methyltetrahydrofolate-binding site of the rat liver methionine

synthase was constructed; docking experiments were designed to elucidate,

in greater detail, the binding mode and reveal structural requirements for

the design of inhibitors of methionine synthase. Our results indicate that

the potency of the tested compounds is related to a planar region of the

inhibitor that can be positioned in the centre of the active site, the presence

of a nitro functional group and two or three probable hydrogen-bonding

interactions.

Abbreviations

CH3-Cbl, methylcobalamin; DHPS, dihydropteroate synthase; Hcy, homocysteine; IC50, half-inhibitory concentration; MeTr, methyltransferase

protein; MetS, methionine synthase; S-AdoMet, S-adenosylmethionine.

FEBS Journal 274 (2007) 287–299 ª 2006 The Authors Journal compilation ª 2006 FEBS 287