Table Of ContentADP-RIBOSYLATION IN
ANIMAL TISSUES
Structure, Function,and Biologyof
Mono (ADP-ribosyl) Transferases
and Related Enzymes
ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY
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NATHAN BACK,State UniversityofNew YorkatBuf/alo
IRUN R. COHEN, The Weizmann InstituteofScience
DAVID KRITCHEVSKY, WistarInstitute
ABEL LAJTHA,N. S. Kline Institutefor PsychiatricResearch
RODOLFO PAOLETTI,UniversityofMilan
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ADP-RIBOSYLATION IN ANIMAL TISSUES: Structure, Function,and BiologyofMono
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ADP-RIBOSYLATION IN
ANIMAL TISSUES
Structure, Function, and Biology of
Mono (ADP-ribosyl) Transferases
and Related Enzymes
Edited by
Friedrich Haag
Friedrich Koch-Nolte
University Hospital Hamburg-Eppendorf
Hamburg. Germany
SPRINGER SCIENCE+BUSINESS MEDIA, LLC
llbrlry of Congresl Cataloging-In-Publication Data
AOP-rl~osylatlon In anl.al tissues structure. function. and biology
of lono (ADP-rlbOSyll transferases anD relateD enZYles I eDiteD by
Friedrich Haag, FriedriCh Koch-Nalte.
p. CI. -- (Advances In exporl •• ntal ledlelne and ~Iology ; v.
419)
-Proceedings of an InternatiOnal Workshop on the BiologIcal
Slgnlflcanc, of Mono ADP-Ribosylatlon In Anl •• l Tissues. helo May
19-23. 1996. In Huburg, Geruny'--Lp. v,rso.
Includes ~Ibllograpl"llcal referencu ind Index.
ISBN 978-1-4613-4652-4 ISBN 978-1-4419-8632-0 (eBook)
DOI 10.1007/978-1-4419-8632-0
I. ADP-rlbosylatlon--Congresses. I. Haag. Friedrich. 11. Koch
-Nolte, FrIedrich. Ill. International HorkshOp on tl'le DialogIcal
Significance of Mono ADP-RI~osylatIOn In Anin i TIssues (1996
H• • burg. Ger.any. lu. Series.
[DNLM: I. Adenosine Diphosphate Rlbos"--.eta~oll'I--congresses.
2. NAD+ ...oP-R I bosy I transferase--.etabo I I u--congresses. HI A0559
v.419 1997 f au 57 A240B 19971
0P625.A29A346 1997
572·.7921--oc21
DNLMIOLC
for Library of Congress 97-7429
Proceedings of an [nternational workshop on the Biological Significance of mono ADP-rioosy[ation in
Anima[ Tissues, held May [9- 23, [996, in Hamburg. Germany
[SBN 978-1-46[3-4652-4
o
1997 Springer Seicnce+Business Media New York
Origina[[y published by Plenum Press, New York in 1997
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PREFACE
Although ADP-ribosylation has been known as a post-translational modification of
proteins for approximately thirty years, the study ofendogenous mono-ADP-ribosylation
in animal tissues has remained somewhat of an orphan field during this time. Until re
cently, interest in the field has concentrated on two types ofphenomena: (I) poly-ADP-ri
bosylation of nuclear proteins in eukaryotes as a mechanism possibly involved in DNA
excision repair and (2) mono-ADP-ribosylation by bacterial enzymes, either as a toxic
mechanism in eukaryotic host cells or as a reversible regulatory mechanism for control of
nitrogen fixation. The identification of diphtheria, cholera, and pertussis toxins as
mono(ADP-ribosyl)transferases and their subsequent purification and crystallization have
shaped ourcurrent knowledge ofthe biology ofmono-ADP-ribosylation reactions and the
structure-function relationships of the enzymes involved. In contrast, endogenous trans
ferases ofanimal tissues escaped molecular cloning, and for a long time their biological
relevance was merely postulated by analogy to theirbacterial cousins.
The molecular characterization of the first mammalian mono(ADP-ribosyl)trans
ferase from rabbit skeletal muscle, as well as the consequent realization that a well-studied
surface membrane protein of T cells, RT6, is a mono(ADP-ribosyl)transferase, have
changed this situation. With their genes at hand, the question ofthe significance ofthese
enzymes in animal tissues can now be addressed more directly. The international work
shop "Biological Significance ofMono-ADP-Ribosylation in Animal Tissues," held May
19-23, 1996, in Hamburg, was the first ofits kind and was felt by many members ofthe
community to be a necessary focal point whose time was now due.
The conference was held in the beautiful setting ofthe Elsa-Brandstrom-Haus, over
looking the shores ofthe Elbe river in Hamburg-Blankenese, and attended by 70 scientists
from Europe, North America, and Japan. It is an interesting historical anecdote that the
first scientific conference to be held at this site was one ofthe early international work
shops on ADP-ribosylation, organized twenty years ago by Helmuth Hilz, Professor
emeritus ofthe Department of Physiological Chemistry and a special guest at this meet
ing. Thirty-nine papers were presented by invited speakers, 22 as posters and short oral
communications. Almost all ofthese are united in this volume, essentially arranged in the
orderthey were presented at the conference.
The first section deals with the lessons to be learned from the studies ofprokaryotic
mono(ADP-ribosyl)transferases. In these enzymes, structure-function relationships have
already been well established by crystallography and site-directed mutagenesis. These
studies facilitate the prediction ofcatalytic residues ofthe eukaryotic transferases, gener
ating hypotheses that can be tested experimentally by site-directed mutagenesis. The regu
lation of the dinitrogenase reductase system in photobacteria also provides an attractive
v
vi Preface
model for reversible regulation ofmetabolic functions by ADP-ribosylation in animal tis
sues.
The second section focuses on the structural and biochemical characterization ofthe
eukaryotic transferases. In the forefront are the reports on the elucidation ofthe primary
structure oftransferases from diverse vertebrate tissues and species. Transferase activity
has been demonstrated also in fungi, although the enzyme(s) involved has/have not yet
been cloned. Some ofthe cloned vertebrate enzymes have already been expressed in a re
combinant form, and mutagenesis experiments have been conducted to test some ofthe
structural hypothesesgenerated in the first section.
Thethirdandfourth sectionsaddress the biological function ofmono-ADP-ribosyla
tion in the immune and othersystems. Several reports sumup what is known to date about
the biological significance of the RT6 T-cell mono(ADP-ribosyl)transferases. RT6 has
been well studied over the years as a T-cell differentiation marker, especially in light of
the observation that RT6 expression defects coincide with increased disease susceptibility
in different animal models for autoimmune diseases. Thus, although its enzymatic activity
has only recently been recognized, RT6 is the transferase whose gene structure has been
the most extensively studied and the system in which hypotheses regarding possible dis
ease association ofdefective transferases are most advanced. ADP-ribosylation reactions
have been implicated in the modulation ofactivation ofnot only T cells, but also other
cells of the immune system, including monocytes/macrophages and granulocytes. In
skeletal and cardiac muscle, target proteins for ADP-ribosylation are being identified,
leading to testable hypotheses as to the biological significance ofthese reactions. Regula
tory effects of ADP-ribosylation have also been demonstrated in central and peripheral
nervous tissue. In the latter case, interestingly, an association between a metabolic disor
der (diabetes mellitus) and endogenous ADP-ribosylation could be found. Finally, exten
sive work has gone into the characterization ofADP-ribosylation reactions in the context
ofregulation ofcellulartransportprocesses.
The fifth section attempts to shed some light on a puzzle that still impedes a clear
understanding ofhow ADP-ribosylation works in animal cells. Although NAD+ is a clas
sic intracellular metabolite, all vertebrate mono(ADP-ribosyl)transferases (and also ADP
ribosyl cyclases) cloned to date are predicted to be localized extracellularly, the majority
as GPI-anchored cell surface enzymes. Therefore, the cellular compartmentalization, dy
namics, and interaction with signal transduction processes of GPI-anchored molecules
havebeenaddressed.
The sixth section examines the mechanisms and biological significance of other
ADP-ribose transfer reactions, especially the production ofcyclic ADPR and related cal
cium-mobilizing secondmessengersbyADP-ribosyl cyclases.
Aspecial session ofthe conference wasdedicated to Prof. Heinz-GunterThiele, one
ofthe pioneers ofresearch on the T-cell mono(ADP-ribosyl)transferase RT6, at the occa
sion ofhis retirement. This session was highlighted by an eloquent pleaby Prof. Jonathan
Howard, anotherone ofthe "fathers"ofRT6, to considerthe implications ofthe extensive
polymorphism within the RT6 system when assessing its function, as this may point to
ward a role ofRT6 within the context ofa defense system that needs to adapt to achang
ingenvironment.
We wish to express our sincere thanks to the Deutsche Forschungsgemeinschaft
(DFG) and the many other groups whose generous financial supportmade this conference
possible, as well as to our students and laboratory staff who worked hard to make the
meetingarewardingexperience for all participants.
Preface vii
We believe that the present volume represents a condensation ofpresent knowledge
ofmono(ADP-ribosyl)transferases and related enzymes and hope that it will fulfill its in
tended function to stimulate and facilitate further research into the significance ofthese
intriguingenzymes.
Friedrich Haag
Friedrich Koch-Nolte
Hamburg. Germany
ACKNOWLEDGMENTS
Financial support for the conference was provided by the following sponsors:
THE DEUTSCHEFORSCHUNGSGEMEINSCHAFT
THE UNIVERSITY HOSPITAL HAMBURG-EPPENDORF
ELIAS
BIOTEST
BOEHRINGER MANNHElM
DIANOVA
DYNAL
SANDOZ
SORVALL
WILKE & WITZEL
BECKMANN
BECTON DICKINSON
BYK SANGTEC
PHARMACIA
SIGMA
SCHLEICHER & SCHUELL
THE BINDING SITE
The organizers thank the department secretary, Ms. Eva-Maria Hobbje; their gradu
ate students Rickmer Braren, Katrin Firner, Martina Matthes, Stefan Rothenburg, and Kar
sten Wursthorn; and their technicians Sandra Bauschus, Roman Girisch, Maren KOhl, and
Cornelia Ritterfor theirsupport.
The organizers thank theirsignificantothers for their understanding.
ix
CONTENTS
INTRODUCTION
I. Mono(ADP-Ribosyl)Transferasesand Related Enzymesin AnimalTissues:
EmergingGene Families .
FriedrichKoch-Nolte andFriedrich Haag
2. ADP-Ribose: AHistoricalOverview 15
Helmuth Hilz
3. ADP-RibosylarginineHydrolasesand ADP-Ribosyltransferases: Partnersin
ADP-RibosylationCycles. ...................................... 25
Joel Moss, AnnaZolkiewska,andIanOkazaki
MONO-ADP-RIBOSYLATION IN PROCARYOTES
PlenaryLectures
4. Crystal StructureofDiphtheriaToxinBoundtoNicotinamideAdenine
Dinucleotide 35
Charles E. Bell and David Eisenberg
5. SelectionofDiphtheriaToxin Active-Site Mutants inYeast: Rediscoveryof
Glutamic Acid-148 asaKey Residue. ............................. 45
HaianFu, Steven R. Blanke,LarryC. Mattheakis, and R. John Collier
6. Identificationofthe Catalytic SiteofClostridial ADP-Ribosyltransferases 53
KlausAktories
7. AProposed Role for Protein: ProteinComplexesin the Regulationofthe
ReversibleADP-RibosylationofDinitrogenaseReductase ... .......... 61
SandraK. Grunwald, YaopingZhang,CaleHalbleib,GaryP. Roberts, and
PaulW. Ludden
8. ADP-Ribosylationand EarlyTranscriptionRegulationbyBacteriophageT4 71
Kai Wilkens, BerndTiemann,FernandoBazan,andWolfgangRuger
xi
xii Contents
PosterReports
9. PertussisToxin: EntryintoCellsandEnzymaticActivity. ................... 83
AlielBayii, RuthLinnemann, LarsvonOlleschik-Elbheim,and
M. AlexanderSchmidt
10. MembraneAnchoredSynthetic PeptidesasaTool forStructure-Function
AnalysisofPertussisToxinand ItsTargetProteins. ................... 87
Lars vonOlleschik-Elbheim,Aliel Bayii,andM. AlexanderSchmidt
II. EnhancedDegradationofStimulatoryG-Protein(Gsa)byCholeraToxinIs
MediatedbyADP-RibosylationofGsa ProteinbutNotbyIncreased
CyclicAMPLevels ............................................. 93
BukhtiarH. Shah
MOLECULARAPPROACHESTOEUCARYOTIC
MONO(ADP-RIBOSYL)TRANSFERASES
PlenaryLectures
12. SequenceandStructuralLinksbetweenDistantADP-Ribosyltransferase
Families. ..................................................... 99
1. FernandoBazanandFriedrichKoch-Nolte
13. MolecularCloningandCharacterizationoftheT-Cell
Mono(ADP-Ribosyl)TransferaseRT6: Relationshipsto OthermADPRTs
andPossibleFunctions .......................... 109
Heinz-GunterThiele,FriedrichHaag,andFriedrichKoch-Nolte
14. MolecularCloningandCharacterizationofaMono(ADP-Ribosyl)Transferase
from HumanTestis 121
IsabelleLevy,Andre Pawlak,MarieGenevieveMattei,andGeorgesGuellaen
15. MolecularCloningandCharacterizationofLymphocyteand Muscle
ADP-Ribosyltransferases 129
Ian1.Okazaki,Hyun-Ju Kim, andJoel Moss
16. MolecularCloningandCharacterizationofArginine-Specific
ADP-Ribosyltransferasesfrom Chicken Bone MarrowCells. ........... 137
MakotoShimoyama,MikakoTsuchiya,NobumasaHara,Kazuo Yamada,and
HarumiOsago
17. Purification,Characterisation,andMolecularCloningofaChickenErythroblast
Mono(ADP-Ribosyl)Transferase 145
TerenceDavis,Jamal S. M. Sabir,ManoochehrTavassoli,andSydneyShall
18. MolecularCharacterisationofaFungalMono(ADP-Ribosyl)Transferase ....... 155
MarthaDeveze-Alvarez,JesusGarcia-Soto,and Guadalupe Martinez-Cadena
