Structure

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A Different Look for AB5 Toxins

F. Xavier Gomis-Ruth1,*1Proteolysis Laboratory,Molecular Biology Institute of BarcelonaCSIC, Barcelona Science Park, Helix Building, c/Baldiri Reixac 15-21, 08028Barcelona, Spain*Correspondence: fxgr@ibmb.csic.eshttp://dx.doi.org/10.1016/j.str.2013.10.004

Metzincins are a distinct clan of metallopeptidases encompassing several families. In this issue of Structure,Ng and colleagues describe the results of the structural analysis of the toxilysins, a novel family of metzincinsemployed by gastroinfective bacteria as intracellular virulence factors following host cell invasion.

Figure 1. Ribbon-type Plot of the A Subunitof AB5 ToxinThe catalytic moiety of the founding member ofthe toxilysin family of metzincin MPs, EcxA, isshown in standard orientation (Gomis-Ruth et al.,2012), i.e., with a view into the active-site cleft.The catalytic metal ion (magenta sphere) is boundby the three histidines (brown sticks) comprisedin the currently revised extended zinc-bindingsignature characteristic for metzincins, HEXXHXX(G/N)XX(H/D). The general base/acid glutamateis in pink, the Met-turn with its methionine sidechain is in blue, the family specific residue is inred, and a disulfide bond, which ligates the up-stream A1 part with the downstream A2 part ofthe A subunit, is depicted in yellow. Characteristicregular secondary-structure elements of metzin-cins are three a helices (in red) and four or five bstrands arranged in a sheet in the N-terminal uppersubdomain (in yellow).

Pathogenic bacteria employ a plethora of

virulence factors to disrupt the function of

infected host cells (Dubin et al., 2013).

Among such factors are the AB5 toxins,

which are complexes between five copies

of a B subunit employed for attachment to

the host plasma membrane and a single

catalytic A subunit, which subverts

cellular functions once the toxin complex

has been internalized (Merritt and Hol,

1995). Four AB5 families have been re-

ported based on the function of the

catalytic subunit: the Shiga toxins, which

interfere with protein biosynthesis; the

pertussis and cholera toxins, which affect

regulation of fluid secretion; and the sub-

tilase cytotoxins, which have a subtilisin-

type serine proteinase as A subunit that

cleaves an endoplasmic reticulum chap-

erone (Beddoe et al., 2010; Le Nours

et al., 2013). More recently, a fifth family

was identified in diarrhea-causing strains

of Escherichia coli and Citrobacter

freundii, represented by toxins EcxAB

and CfxAB, respectively (Jansson et al.,

2010; Karasawa et al., 2002).

In this issue ofStructure, Ng et al. (2013)

present the crystal structure of the E. coli

EcxAB toxin complex, which allows them

to determine the molecular determinants

of function of this heterohexameric viru-

lence factor. Similar to previous AB5

complex structures, the B subunits are

arranged in a ring around a central pore

aligned with a 5-fold axis. The A subunit

sits on top of this ring and introduces its

C-terminal helical tail into the pore of the

B pentamer. The authors reveal that the

catalytic EcxA subunit is a metallopepti-

dase (MP) that is unrelated to any other

AB5 family. It presents structural similarity

with matrix metalloproteinases, which are

a family within the metzincin clan of MPs.

However, EcxA has distinct extra struc-

tural elements that give rise to a fold

different from any metzincin structurally

characterized to date (Gomis-Ruth,

2003, 2009). Accordingly, the authors

propose the name toxilysins to refer to a

new family comprising EcxA and several

CfxA-type proteins.

Metzincins were discovered back in

1993 by Bode et al. (1993), acknowl-

edging structural similarities among three

MP prototypes that had been structurally

characterized by then, namely crayfish

astacin (astacin family), snake-venom

adamalysin II (adamalysin/ADAM family),

and Pseudomonas aeruginosa alkaline

proteinase (serralysin family) (Stocker

et al., 1995). Despite a negligible

sequence identity and distinct lengths of

their polypeptide chains, these enzymes

showed topologically equivalent struc-

tural features: subdivison of the catalytic

moieties into an upper N-terminal subdo-

main (NTS) and a lower C-terminal sub-

domain (CTS) by an active-site cleft

harboring the catalytic metal ion at its

bottom; two helices termed the ‘‘backing

helix’’ and the ‘‘active-site helix’’ within

the NTS; a five-stranded mixed parallel/

antiparallel b sheet, likewise in the NTS,

whose lowermost strand created the

upper rim of the active-site cleft; a ‘‘C-ter-

minal helix’’ at the end of the CTS; and,

most noteworthy, a strictly conserved

1,4 turn immediately below the metal

site featuring a methionine in its third

position, the ‘‘Met-turn.’’ In addition, the

active-site helix and the subsequent

part of the polypeptide included the ex-

tended zinc-binding consensus sequence,

HEXXHXXGXXH, with the three histidines

liganding the catalytic metal ion, the gluta-

mate acting as a general base/acid during

catalysis, and the glycine accounting for a

characteristic sharpchange in thedirection

of the polypeptide after the active-site helix

to reach the third histidine. Thereafter,

Structure 21, November 5, 2013 ª

the catalytic domains of other MPs were

successively reported. All featured the

aforementioned common structural traits

despite minute sequence similarity as well

as family-specific structural elements,

such as additional ion-binding sites, in-

serted domains, and upstream and down-

stream polypeptide extensions that

caused them to span between �130 and

�270 residues. These MPs became the

founding members of novel families within

the metzincins: matrix metalloproteinases

or matrixins (reported for their structure

for the first time in 1994), leishmanolysins

2013 Elsevier Ltd All rights reserved 1909

Structure

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(1995), snapalysins (1997), pappalysins

(2006), archaemetzincins (2010), fragily-

sins (2010), igalysins (deposited with the

ProteinDataBank in2011, thoughnot pub-

lished), and cholerilysins (2012). The pre-

sent work reveals that EcxA comprises all

the characteristic elements of metzincins

(Figure 1), and we welcome the founding

member of a new metzincin family, the

toxilysins.

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