A. Corti

A. Pompella

FRANZINI, Maria

Frontiers in Pharmacology

PubMed

Maria Franzini

Alfonso Pompella

Alessandro Corti

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Gamma-glutamyltransferases: exploring the complexity of a multi-functional family of enzymes

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Archivio della ricerca della Scuola Superiore Sant'Anna

BOOK REVIEWpublished: 30 September 2014doi: 10.3389/fphar.2014.00221Gamma-glutamyltransferases: exploring the complexity ofa multi-functional family of enzymesMaria Franzini1,2, Alfonso Pompella3 and Alessandro Corti3*1 Cardiology and Cardiovascular Medicine Division, Fondazione Toscana G. Monasterio – CNR, Pisa, Italy2 Institute of Life Sciences, Scuola Superiore Sant’Anna, Pisa, Italy3 Department of Translational Research and New Technologies in Medicine and Surgery, Medical School, University of Pisa, Pisa, Italy*Correspondence: alessandro.corti@med.unipi.itEdited by:Chiranjib Chakraborty, Galgotias University, IndiaReviewed by:Salvatore Salomone, Catania University, ItalyKeywords: gamma-glutamyltransferase, glutathione, gamma-glutamyltransferase structure, gamma-glutamyltransferase fractions, microvesicles,biotechnological applications, prodrugsA book review onGamma-Glutamyl Transpeptidases: Structure and FunctionEdited by Castellano, I., and Merlino, A., Heidelberg: Springer, 2013.Gamma-glutamyltransferase (γ-GT; EC2.3.2.2) is the sole enzyme activity capa-ble of cleaving the gamma-glutamyl bondpresent in the structure of glutathione(GSH), the well known antioxidant tripep-tide, and is present on the outer aspect ofplasma membranes of virtually all mam-malian cells. γ-GT–mediated cleavage isthe first step in the degradation of extra-cellular GSH, which proceeds by the actionof dipeptidases and makes thus precur-sor aminoacids (cysteine in the first place)available for reuptake and re-utilization bythe cell for GSH resynthesis. On the otherhand, γ-GT is also responsible for the pro-cessing of other substrates including GSHin their structure, such as major inflam-matory mediators leukotriene C4 (LTC4)and S-nitroso-glutathione (GSNO), aswell as adducts of GSH with xenobioticsand metabolites formed by the action ofglutathione-S-transferases.Due to its critical role in cellular GSHresupply, γ-GT has long been consid-ered as just a part of cellular antioxidantdefenses. However, prooxidant effects ofγ-GT were also documented (Dominiciet al., 2005), and the significance ofthis enzyme activity in cellular phys-iology and medicine has considerablyexpanded as its central role in modula-tion of overall redox equilibria of the cellhas become increasingly clear. In theirbook “Gamma-Glutamyl Transpeptidases:Structure and Function” (Castellano andMerlino, 2013) the authors provide a com-prehensive coverage of the state-of-artknowledge on many relevant aspects of thematter. After a brief general introduction,Chapter 2 provides an updated overviewof the complex γ-GT gene structure, aswell as of expression, induction and dis-tribution of the enzyme in mammaliantissues. The following chapters are con-cerned with structure-function correlates,and start with an interesting section onthe enzyme “autoprocessing,” i.e., the pro-cess by which precursor γ-GT protein—translated as a single polypeptide—exertsa proteolytic cleavage on its own structurethus yielding a large and a small subunitswhich then assemble together to give themature enzyme dimer. To date structuralcharacterization of human γ-GT1 (hGT1)has been hampered by its heavily glyco-sylated structure, as well as by its associ-ation with cell membranes, and most ofthe knowledge regarding enzyme struc-ture indeed comes from investigations inbacteria. The text analyzes in detail thecharacteristics of enzyme active site andfactors affecting substrate binding affin-ity, the reaction mechanism and its steps,the mechanisms of enzyme inhibition byseveral distinct compounds, the assaysavailable to measure enzyme activity, thebiochemical characteristics of the enzymeprotein, etc. This part—Chapters 3 and4—form the core of the book, in which theauthors directly capitalize the competencesdeveloped in their own laboratory.Nevertheless, the text is forcedly lack-ing an important aspect of the matter, i.e.the recently published characterization ofmacromolecular complexes carrying γ-GTin biological fluids. It has been in fact doc-umented that γ-GT associates in plasmawith macromolecular carriers having dif-ferent molecular weights, densities andcharges, and recently four distinct γ-GTfractions were identified (Franzini et al.,2013). A first characterization has shownthat the “b-GGT” fraction consists ofmembrane microvesicles resembling exo-somes (Fornaciari et al., 2014). Additionalinsights into hGT1 high-resolution crystalstructure have also been recently reported(West et al., 2013), highlighting specificdifferences between bacterial and humanarchitectures.A better knowledge of γ-GT struc-ture will undoubtly help to clarify itsinteractions with carriers and improve itsclinical value and specificity as a dis-ease biomarker. Chapter 5 (“γ-GT inBioclinical”) indeed provides an overviewof the pathophysiological roles playedby γ-GT in human pathology. In clin-ical medicine, γ-GT plasma levels arewww.frontiersin.org September 2014 | Volume 5 | Article 221 | 1Franzini et al. Multifunctionality of gamma-glutamyltransferasesconsidered a well established and sensi-tive index of hepato-biliary disease andalcohol abuse, but during the last decadethe involvement of γ-GT in other impor-tant diseases has also been documented.A series of large-scale, prospective stud-ies have shown that plasma γ-GT valuesare independent predictors of cardiovascu-lar mortality, stroke, metabolic syndromeand diabetes, which appears to confirm theinvolvement of γ-GT in mechanisms offlogosis and cellular damage. Expression ofγ-GT during cancer progression is a factorin drug resistance of cancer cells, e.g., tocisplatin. This part however is a less inter-esting section of the book, as the same top-ics were previously discussed in detail byothers (Pompella et al., 2007; Corti et al.,2010).The final Chapter 6 includes an accu-rate and compelling overview to thebiotechnological and biomedical appli-cations of γ-GT, exploiting the uniquecapability of this enzyme to cleave thegamma-glutamyl bond. For this reason,e.g., the addition of a gamma-glutamylgroup to a drug molecule can make itinactive until the moment the residueis cleaved off by γ-GT. The examplesdescribed include the synthesis of gamma-D-glutamyl-L-tryptophan (a compoundemployed for treatment of tuberculo-sis) and phenylhydrazine derivatives,exhibiting antioxidant and immunos-timulatory properties. The use of γ-GTfrom Bacillus subtilis—which can act ascephalosporin acylase—in the produc-tion of cephalosporins, and the possibleuse of γ-GT in the production of thea-nine are also discussed. The Authorsmight have as well cited the recentdevelopment of the GSH-containingarsenic trioxide–based pro-drug 4-(N-(S-glutathionylacetyl)amino)phenylarsinousacid (GSAO), which is selectively activatedby γ-GT expressed by cancer cells andexerts a remarkable anti-angiogenic effect(Dilda et al., 2008).In summary, the book by Castellanoand Merlino covers a full spectrum of rele-vant topics regarding γ-GT biochemistry,pathophysiology and biotechnology. Thisconcise text—integrated by a combinationof comprehensive and useful tables, fig-ures, and graphs reporting essential infor-mation in a clear and detailed way—can provide a precious introduction toγ-GT, a complex and rapidly developingfield of interest for a wide readership ofbiochemists, biologists, and clinicians.REFERENCESCastellano, I., and Merlino, A. (2013). Gamma-Glutamyl Transpeptidases: Structure and Function.Heidelberg: Springer. doi: 10.1007/978-3-0348-0682-4Corti, A., Franzini, M., Paolicchi, A., and Pompella,A. (2010). Gamma-glutamyltransferase of can-cer cells at the crossroads of tumor progression,drug resistance and drug targeting. Anticancer Res.30, 1169–1181. Available online at: http://ar.iiarjournals.org/content/30/4/1169.full.pdfDilda, P. J., Ramsey, E. E., Corti, A., Pompella,A., and Hogg, P. J. (2008). Metabolism ofthe tumor angiogenesis inhibitor 4-(N-(S-glutathionylacetyl)amino)phenylarsinous acid.J. Biol. Chem. 283, 35428–35434. doi: 10.1074/jbc.M804470200Dominici, S., Paolicchi, A., Corti, A., Maellaro,E., and Pompella, A. (2005). Prooxidant reac-tions promoted by soluble and cell-boundγ-glutamyltransferase activity. Methods Enzymol.401, 483–500. doi: 10.1016/S0076-6879(05)01029-3Fornaciari, I., Fierabracci, V., Corti, A., Aziz Elawadi,H., Lorenzini, E., Emdin, M., et al. (2014).Gamma- glutamyltransferase fractions in humanplasma and bile: characteristic and biogene-sis. PLoS ONE 9:e88532. doi: 10.1371/jour-nal.pone.0088532Franzini, M., Fornaciari, I., Rong, J., Larson,M. G., Passino, C., Emdin, M., et al. (2013).Correlates and reference limits of plasmagamma-glutamyltransferase fractions fromthe Framingham Heart Study. Clin. Chim. Acta417, 19–25. doi: 10.1016/j.cca.2012.12.002Pompella, A., Corti, A., Paolicchi, A., Giommarelli,C., and Zunino, F. (2007). Gamma-glutamyltransferase, redox regulation and cancerdrug resistance. Curr. Opin. Pharmacol. 7,360–366. doi: 10.1016/j.coph.2007.04.004West, M. B., Chen, Y., Wickham, S., Heroux, A.,Cahill, K., Hanigan, M. H., et al. (2013). Novelinsights into eukaryotic γ-glutamyltranspeptidase1 from the crystal structure of the glutamate-bound human enzyme. J. Biol. Chem.288, 31902–31913. doi: 10.1074/jbc.M113.498139Conflict of Interest Statement: The authors declarethat the research was conducted in the absence of anycommercial or financial relationships that could beconstrued as a potential conflict of interest.Received: 09 June 2014; accepted: 12 September 2014;published online: 30 September 2014.Citation: Franzini M, Pompella A and Corti A (2014)Gamma-glutamyltransferases: exploring the complex-ity of a multi-functional family of enzymes. Front.Pharmacol. 5:221. doi: 10.3389/fphar.2014.00221This article was submitted to ExperimentalPharmacology and Drug Discovery, a section ofthe journal Frontiers in Pharmacology.Copyright © 2014 Franzini, Pompella and Corti. Thisis an open-access article distributed under the terms ofthe Creative Commons Attribution License (CC BY).The use, distribution or reproduction in other forumsis permitted, provided the original author(s) or licen-sor are credited and that the original publication inthis journal is cited, in accordance with accepted aca-demic practice. No use, distribution or reproduction ispermitted which does not comply with these terms.Frontiers in Pharmacology | Experimental Pharmacology and Drug Discovery September 2014 | Volume 5 | Article 221 | 2

Gamma-glutamyltransferases: exploring the complexity of a multifunctional family of enzymes

FRANZINI, MARIA

POMPELLA, ALFONSO

CORTI, ALESSANDRO

Archivio della Ricerca - Università di Pisa

BOOK REVIEW
published: 30 September 2014
doi: 10.3389/fphar.2014.00221
Gamma-glutamyltransferases: exploring the complexity of
a multi-functional family of enzymes
Maria Franzini1,2, Alfonso Pompella3 and Alessandro Corti3*
1 Cardiology and Cardiovascular Medicine Division, Fondazione Toscana G. Monasterio – CNR, Pisa, Italy
2 Institute of Life Sciences, Scuola Superiore Sant’Anna, Pisa, Italy
3 Department of Translational Research and New Technologies in Medicine and Surgery, Medical School, University of Pisa, Pisa, Italy
*Correspondence: alessandro.corti@med.unipi.it
Edited by:
Chiranjib Chakraborty, Galgotias University, India
Reviewed by:
Salvatore Salomone, Catania University, Italy
Keywords: gamma-glutamyltransferase, glutathione, gamma-glutamyltransferase structure, gamma-glutamyltransferase fractions, microvesicles,
biotechnological applications, prodrugs
A book review on
Gamma-Glutamyl Transpeptidases: Structure and Function
Edited by Castellano, I., and Merlino, A., Heidelberg: Springer, 2013.
Gamma-glutamyltransferase (γ-GT; EC
2.3.2.2) is the sole enzyme activity capa-
ble of cleaving the gamma-glutamyl bond
present in the structure of glutathione
(GSH), the well known antioxidant tripep-
tide, and is present on the outer aspect of
plasma membranes of virtually all mam-
malian cells. γ-GT–mediated cleavage is
the first step in the degradation of extra-
cellular GSH, which proceeds by the action
of dipeptidases and makes thus precur-
sor aminoacids (cysteine in the first place)
available for reuptake and re-utilization by
the cell for GSH resynthesis. On the other
hand, γ-GT is also responsible for the pro-
cessing of other substrates including GSH
in their structure, such as major inflam-
matory mediators leukotriene C4 (LTC4)
and S-nitroso-glutathione (GSNO), as
well as adducts of GSH with xenobiotics
and metabolites formed by the action of
glutathione-S-transferases.
Due to its critical role in cellular GSH
resupply, γ-GT has long been consid-
ered as just a part of cellular antioxidant
defenses. However, prooxidant effects of
γ-GT were also documented (Dominici
et al., 2005), and the significance of
this enzyme activity in cellular phys-
iology and medicine has considerably
expanded as its central role in modula-
tion of overall redox equilibria of the cell
has become increasingly clear. In their
book “Gamma-Glutamyl Transpeptidases:
Structure and Function” (Castellano and
Merlino, 2013) the authors provide a com-
prehensive coverage of the state-of-art
knowledge on many relevant aspects of the
matter. After a brief general introduction,
Chapter 2 provides an updated overview
of the complex γ-GT gene structure, as
well as of expression, induction and dis-
tribution of the enzyme in mammalian
tissues. The following chapters are con-
cerned with structure-function correlates,
and start with an interesting section on
the enzyme “autoprocessing,” i.e., the pro-
cess by which precursor γ-GT protein—
translated as a single polypeptide—exerts
a proteolytic cleavage on its own structure
thus yielding a large and a small subunits
which then assemble together to give the
mature enzyme dimer. To date structural
characterization of human γ-GT1 (hGT1)
has been hampered by its heavily glyco-
sylated structure, as well as by its associ-
ation with cell membranes, and most of
the knowledge regarding enzyme struc-
ture indeed comes from investigations in
bacteria. The text analyzes in detail the
characteristics of enzyme active site and
factors affecting substrate binding affin-
ity, the reaction mechanism and its steps,
the mechanisms of enzyme inhibition by
several distinct compounds, the assays
available to measure enzyme activity, the
biochemical characteristics of the enzyme
protein, etc. This part—Chapters 3 and
4—form the core of the book, in which the
authors directly capitalize the competences
developed in their own laboratory.
Nevertheless, the text is forcedly lack-
ing an important aspect of the matter, i.e.
the recently published characterization of
macromolecular complexes carrying γ-GT
in biological fluids. It has been in fact doc-
umented that γ-GT associates in plasma
with macromolecular carriers having dif-
ferent molecular weights, densities and
charges, and recently four distinct γ-GT
fractions were identified (Franzini et al.,
2013). A first characterization has shown
that the “b-GGT” fraction consists of
membrane microvesicles resembling exo-
somes (Fornaciari et al., 2014). Additional
insights into hGT1 high-resolution crystal
structure have also been recently reported
(West et al., 2013), highlighting specific
differences between bacterial and human
architectures.
A better knowledge of γ-GT struc-
ture will undoubtly help to clarify its
interactions with carriers and improve its
clinical value and specificity as a dis-
ease biomarker. Chapter 5 (“γ-GT in
Bioclinical”) indeed provides an overview
of the pathophysiological roles played
by γ-GT in human pathology. In clin-
ical medicine, γ-GT plasma levels are
www.frontiersin.org September 2014 | Volume 5 | Article 221 | 1
Franzini et al. Multifunctionality of gamma-glutamyltransferases
considered a well established and sensi-
tive index of hepato-biliary disease and
alcohol abuse, but during the last decade
the involvement of γ-GT in other impor-
tant diseases has also been documented.
A series of large-scale, prospective stud-
ies have shown that plasma γ-GT values
are independent predictors of cardiovascu-
lar mortality, stroke, metabolic syndrome
and diabetes, which appears to confirm the
involvement of γ-GT in mechanisms of
flogosis and cellular damage. Expression of
γ-GT during cancer progression is a factor
in drug resistance of cancer cells, e.g., to
cisplatin. This part however is a less inter-
esting section of the book, as the same top-
ics were previously discussed in detail by
others (Pompella et al., 2007; Corti et al.,
2010).
The final Chapter 6 includes an accu-
rate and compelling overview to the
biotechnological and biomedical appli-
cations of γ-GT, exploiting the unique
capability of this enzyme to cleave the
gamma-glutamyl bond. For this reason,
e.g., the addition of a gamma-glutamyl
group to a drug molecule can make it
inactive until the moment the residue
is cleaved off by γ-GT. The examples
described include the synthesis of gamma-
D-glutamyl-L-tryptophan (a compound
employed for treatment of tuberculo-
sis) and phenylhydrazine derivatives,
exhibiting antioxidant and immunos-
timulatory properties. The use of γ-GT
from Bacillus subtilis—which can act as
cephalosporin acylase—in the produc-
tion of cephalosporins, and the possible
use of γ-GT in the production of thea-
nine are also discussed. The Authors
might have as well cited the recent
development of the GSH-containing
arsenic trioxide–based pro-drug 4-(N-(S-
glutathionylacetyl)amino)phenylarsinous
acid (GSAO), which is selectively activated
by γ-GT expressed by cancer cells and
exerts a remarkable anti-angiogenic effect
(Dilda et al., 2008).
In summary, the book by Castellano
and Merlino covers a full spectrum of rele-
vant topics regarding γ-GT biochemistry,
pathophysiology and biotechnology. This
concise text—integrated by a combination
of comprehensive and useful tables, fig-
ures, and graphs reporting essential infor-
mation in a clear and detailed way—
can provide a precious introduction to
γ-GT, a complex and rapidly developing
field of interest for a wide readership of
biochemists, biologists, and clinicians.
REFERENCES
Castellano, I., and Merlino, A. (2013). Gamma-
Glutamyl Transpeptidases: Structure and Function.
Heidelberg: Springer. doi: 10.1007/978-3-0348-
0682-4
Corti, A., Franzini, M., Paolicchi, A., and Pompella,
A. (2010). Gamma-glutamyltransferase of can-
cer cells at the crossroads of tumor progression,
drug resistance and drug targeting. Anticancer Res.
30, 1169–1181. Available online at: http://ar.iiar
journals.org/content/30/4/1169.full.pdf
Dilda, P. J., Ramsey, E. E., Corti, A., Pompella,
A., and Hogg, P. J. (2008). Metabolism of
the tumor angiogenesis inhibitor 4-(N-(S-
glutathionylacetyl)amino)phenylarsinous acid.
J. Biol. Chem. 283, 35428–35434. doi: 10.
1074/jbc.M804470200
Dominici, S., Paolicchi, A., Corti, A., Maellaro,
E., and Pompella, A. (2005). Prooxidant reac-
tions promoted by soluble and cell-bound
γ-glutamyltransferase activity. Methods Enzymol.
401, 483–500. doi: 10.1016/S0076-6879(05)
01029-3
Fornaciari, I., Fierabracci, V., Corti, A., Aziz Elawadi,
H., Lorenzini, E., Emdin, M., et al. (2014).
Gamma- glutamyltransferase fractions in human
plasma and bile: characteristic and biogene-
sis. PLoS ONE 9:e88532. doi: 10.1371/jour-
nal.pone.0088532
Franzini, M., Fornaciari, I., Rong, J., Larson,
M. G., Passino, C., Emdin, M., et al. (2013).
Correlates and reference limits of plasma
gamma-glutamyltransferase fractions from
the Framingham Heart Study. Clin. Chim. Acta
417, 19–25. doi: 10.1016/j.cca.2012.12.002
Pompella, A., Corti, A., Paolicchi, A., Giommarelli,
C., and Zunino, F. (2007). Gamma-
glutamyltransferase, redox regulation and cancer
drug resistance. Curr. Opin. Pharmacol. 7,
360–366. doi: 10.1016/j.coph.2007.04.004
West, M. B., Chen, Y., Wickham, S., Heroux, A.,
Cahill, K., Hanigan, M. H., et al. (2013). Novel
insights into eukaryotic γ-glutamyltranspeptidase
1 from the crystal structure of the glutamate-
bound human enzyme. J. Biol. Chem.
288, 31902–31913. doi: 10.1074/jbc.M113.4
98139
Conflict of Interest Statement: The authors declare
that the research was conducted in the absence of any
commercial or financial relationships that could be
construed as a potential conflict of interest.
Received: 09 June 2014; accepted: 12 September 2014;
published online: 30 September 2014.
Citation: Franzini M, Pompella A and Corti A (2014)
Gamma-glutamyltransferases: exploring the complex-
ity of a multi-functional family of enzymes. Front.
Pharmacol. 5:221. doi: 10.3389/fphar.2014.00221
This article was submitted to Experimental
Pharmacology and Drug Discovery, a section of
the journal Frontiers in Pharmacology.
Copyright © 2014 Franzini, Pompella and Corti. This
is an open-access article distributed under the terms of
the Creative Commons Attribution License (CC BY).
The use, distribution or reproduction in other forums
is permitted, provided the original author(s) or licen-
sor are credited and that the original publication in
this journal is cited, in accordance with accepted aca-
demic practice. No use, distribution or reproduction is
permitted which does not comply with these terms.
Frontiers in Pharmacology | Experimental Pharmacology and Drug Discovery September 2014 | Volume 5 | Article 221 | 2


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Gamma-glutamyltransferases: exploring the complexity of a multifunctional family of enzymes

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