Tài liệu Báo cáo khoa học: Unraveling the catalytic mechanism of lactoperoxidase and myeloperoxidase

REVIEW ARTICLE

Unraveling the catalytic mechanism of lactoperoxidase and

myeloperoxidase

A reflection on some controversial features

Elena Ghibaudi and Enzo Laurenti

Dipartimento di Chimica I.F.M., Universita` di Torino, Italy

Although belonging to the widely investigated peroxidase

superfamily, lactoperoxidase (LPO) and myeloperoxidase

(MPO) share structural and functional features that make

them peculiar with respect to other enzymes of the same

group. A survey of the available literature on their catalytic

intermediates enabled us to ask some questions that

remained unanswered. These questions concern controver￾sial features of the LPO and MPO catalytic cycle, such as the

existence of Compound I and Compound II isomers and

the identification of their spectroscopic properties. After

addressing each of these questions, we formulated a hypo￾thesis that describes an integrated vision of the catalytic

mechanism of both enzymes. The main points are: (a) a

re-evaluation of the role of superoxide as a reductant in the

catalytic cycle; (b) the existence of Cpd I isomers; (c) reci￾procal interactions between catalytic intermediates and (d)

a mechanistic explanation for catalase activity in both

enzymes.

Keywords: lactoperoxidase; myeloperoxidase; aminoacid

radical; Compound I; Compound II; Compound III;

catalytic intermediates.

Introduction

The catalytic cycle of peroxidases, including lactoperoxidase

(LPO) and myeloperoxidase (MPO), is described usually as

a sequence of three consecutive reactions, according to

Scheme 1.

Compound I (Cpd I), which arises from the reaction of

the native enzyme with hydrogen peroxide (H2O2), is two

oxidizing equivalents above the resting state. It reacts with

a substrate molecule and is converted into a secondary

compound that has lost one equivalent, generally indicated

as Compound II (Cpd II). A second substrate molecule

recycles Cpd II into the resting enzyme. A large excess of

H2O2 converts Cpd I into the inactive intermediate, Com￾pound III (Cpd III). The two oxidizing equivalents of

Cpd I are on an iron ion, that assumes the formal oxidation

state IV, and on the porphyrin ring, which becomes a

cationic radical. Cpd II has been shown to contain FeIV¼O

[1–3], whereas Cpd III is an enzyme adduct with superoxide

[2–6]. Depending on the type of peroxidase, Cpd III

formation may be reversible, whereby it can be reconverted

into an active form of the enzyme, or irreversible, in which

case it is associated with degradation of the enzyme.

Moreover, a few peroxidases, e.g. haloperoxidases, can

oxidize halides through the bielectronic reduction of Cpd I

that is converted back to the resting state without forming

Cpd II [3,7,8].

The generally accepted definition of the three intermedi￾ates of this class of enzymes can be misleading. In fact, when

comparing different peroxidases, the same name is applied

to species with distinct electronic structures. Moreover,

several peroxidase intermediates are known where the

unpaired electron is localized onto an amino acid of the

protein scaffold [9–14] and this aspect is not taken into

account by the classical peroxidase cycle.

Within this context, we propose to re-examine some of

the literature data describing the catalytic cycle of two

mammalian peroxidases, LPO and MPO, in order to

reconcile the apparent inconsistencies and to provide some

new insights. MPO and LPO share functional and

structural homology, reflecting their common phylogenetic

origin [15] and participate in antimicrobial host defense,

generating potent reactive species by the oxidation of

halides or pseudohalides. Based on our survey of the

experimental data concerning the reactivity of these

enzymes, we formulated four questions that are focused

on controversial features of the LPO and/or MPO

catalytic cycle: (a) is formation of Cpd I reversible (or

do mammalian peroxidases possess catalase activity); (b)

does Cpd I exist in two isomericforms, containing the

porphyrin radical and the amino acid radical (aa+•

),

respectively; (c) as the conversion of Cpd I fi Cpd II

occurs spontaneously in the presence of peroxide, which is

the reducing agent in this reaction step and (d) are the

optical spectra of Cpd I–[FeIV¼O; aa+•

] and Cpd II

identical?

Correspondence to E. Ghibaudi, Dipartimento di Chimica I.F.M.,

Universita` di Torino, Via Giuria 7 – 10125 Torino, Italy.

Fax: + 39 011 670 7855, Tel.: + 39 011 670 7951,

E-mail: [email protected]

Abbreviations: LPO, lactoperoxidase; MPO, myeloperoxidase;

Cpd I–III, Compound I–III.

Enzymes: lactoperoxidase (EC 1.11.1.7); myeloperoxidase

(EC 1.11.1.7).

(Received 10 April 2003, revised 18 July 2003,

accepted 23 September 2003)

Eur. J. Biochem. 270, 4403–4412 (2003) FEBS 2003 doi:10.1046/j.1432-1033.2003.03849.x