Amino Acids, Peptides and Proteins
in Organic Chemistry

Edited by
Andrew B. Hughes

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Further Reading
Fessner, W.-D., Anthonsen, T.

Budisa, N.

Modern Biocatalysis

Engineering the Genetic Code

Stereoselective and Environmentally
Friendly Reactions

Expanding the Amino Acid Repertoire
for the Design of Novel Proteins

2009

2006

ISBN: 978-3-527-32071-4

ISBN: 978-3-527-31243-6

Sewald, N., Jakubke, H.-D.

Demchenko, A. V. (ed.)

Peptides: Chemistry and
Biology

Handbook of Chemical
Glycosylation

2009

Advances in Stereoselectivity and
Therapeutic Relevance

ISBN: 978-3-527-31867-4

Lutz, S., Bornscheuer, U. T. (eds.)

2008
ISBN: 978-3-527-31780-6

Protein Engineering
Handbook

Lindhorst, T. K.

2 Volume Set

2009

Essentials of Carbohydrate
Chemistry and Biochemistry

ISBN: 978-3-527-31850-6

2007
ISBN: 978-3-527-31528-4

Aehle, W. (ed.)

Enzymes in Industry
Production and Applications
2007
ISBN: 978-3-527-31689-2

Wiley-VCH (ed.)

Ullmann’s Biotechnology and
Biochemical Engineering
2 Volume Set
2007
ISBN: 978-3-527-31603-8

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Amino Acids, Peptides and Proteins
in Organic Chemistry

Volume 2 - Modified Amino Acids, Organocatalysis
and Enzymes

Edited by
Andrew B. Hughes

www.pdfgrip.com


The Editor
Andrew B. Hughes
La Trobe University
Department of Chemistry
Victoria 3086
Australia

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produced. Nevertheless, authors, editors, and
publisher do not warrant the information contained
in these books, including this book, to be free of
errors. Readers are advised to keep in mind that
statements, data, illustrations, procedural details or
other items may inadvertently be inaccurate.
Library of Congress Card No.: applied for
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A catalogue record for this book is available from the
British Library.
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the Deutsche Nationalbibliothek
The Deutsche Nationalbibliothek lists this

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detailed bibliographic data are available in the
Internet at .
# 2009 WILEY-VCH Verlag GmbH & Co. KGaA,
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All rights reserved (including those of translation into
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Printed on acid-free paper
ISBN: 978-3-527-32098-1

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V

Contents
List of Contributors


XIX

Part One Synthesis and Chemistry of Modified Amino Acids
1
1.1
1.2
1.2.1
1.2.1.1
1.2.1.1.1
1.2.1.1.2
1.2.1.2
1.2.1.3
1.2.1.4
1.2.2
1.2.2.1
1.2.2.2
1.2.2.3
1.2.3
1.2.3.1
1.2.3.2
1.3
1.3.1
1.3.1.1
1.3.1.2
1.3.1.3
1.3.1.3.1

1

Synthesis and Chemistry of a,b-Didehydroamino Acids 3

Uli Kazmaier
Introduction 3
Synthesis of DDAAs 3
DDAAs via Eliminations 3
DDAAs via b-Elimination 3
From b-Hydroxy Amino Acids 3
From b-Thio- and Selenoamino Acids 5
Elimination from N-Hydroxylated and -Chlorinated Amino Acids
and Peptides 6
DDAAs from a-Oxo Acids and Amides 6
DDAAs from Azides 7
DDAAs via C¼C Bond Formation 7
DDAAs via Azlactones [5(4H)-Oxazolones] 7
DDAAs via Horner–Emmons and Wittig Reactions 8
DDAAs via Enolates of Nitro- and Isocyano- and Iminoacetates 10
DDAAs via C–C Bond Formation 12
DDAAs via Heck Reaction 12
DDAAs via Cross-Coupling Reactions 13
Reactions of DDAAs 14
Additions to the C¼C Bond 14
Nucleophilic Additions 14
Radical Additions 15
Cycloadditions 17
[3ỵ2] Cycloadditions 18

Amino Acids, Peptides and Proteins in Organic Chemistry.
Vol.2 – Modified Amino Acids, Organocatalysis and Enzymes. Edited by Andrew B. Hughes
Copyright Ó 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
ISBN: 978-3-527-32098-1


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VI

Contents

1.3.1.3.2
1.3.1.4
1.3.2
1.4
1.5
1.5.1
1.5.2
1.5.3
1.5.4
1.5.5
1.5.6
1.5.7
1.5.8

2
2.1
2.1.1
2.1.2
2.1.3
2.1.4
2.2
2.2.1
2.2.1.1

2.2.1.2
2.2.1.3
2.2.1.4
2.2.1.5
2.2.2
2.2.2.1

[4ỵ2] Cycloadditions 19
Catalytic Hydrogenations 19
Halogenations of DDAAs 21
Conclusions 21
Experimental Procedures 22
General Procedure for the Two-Step Synthesis of Dehydroisoleucine
Derivatives 22
General Procedure for the Synthesis of a,b-Didehydroamino Acid Esters
by the Phosphorylglycine Ester Method using DBU 22
General Procedure for the Synthesis of a-Chloroglycine Derivatives 23
General Procedure for the Synthesis of Homomeric Dimers 23
General Procedure for the Synthesis of (Z)-g-Alkyl-a,
b-Didehydroglutamates from Imino Glycinates 24
Palladium-Catalyzed Trifold Heck Coupling 25
General Experimental Procedure for Conjugate Addition of Alkyl
iodides to Chiral a,b-Unsaturated Amino Acid Derivatives 25
Bromination of N-tert-Butyloxycarbonyldehydroamino Acids 26
References 26
Synthesis and Chemistry of a-Hydrazino Acids 35
Joëlle Vidal
Introduction 35
a-Hydrazino Acids are Potent Inhibitors of Pyridoxal
Phosphate Enzymes 35

Natural Products Containing the N–N–C–C¼O Fragment 36
Synthetic Bioactive Products Containing the N–N–C–C¼O
Fragment 39
The CO–N–N–C–CO–NH Fragment is a Turn Inducer in
Pseudopeptides 40
Synthesis 41
Disconnection 1a: Reaction of Hydrazine Derivatives with
Carbon Electrophiles 41
Reaction of Hydrazine Derivatives with Enantiopure
a-Halogeno Acids 42
Reaction of Hydrazine Derivatives with Enantiopure Activated
a-Hydroxy Esters 42
Mitsunobu Reaction of Aminophthalimide Derivatives with
Enantiopure a-Hydroxy Esters 43
Reaction of Hydrazine Derivatives with Nonracemic Epoxides 43
Enantioselective Conjugate Addition of Hydrazines to
a,b-Unsaturated Imides 44
Disconnection 1b: Stereoselective Synthesis using
Azodicarboxylates 44
Stereoselective a-Hydrazination of Chiral Carbonyl Compounds
using Azodicarboxylates 45

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Contents

2.2.2.2
2.2.2.3
2.2.3

2.2.3.1
2.2.3.2
2.2.3.3
2.2.3.4
2.2.4
2.2.4.1
2.2.4.2
2.2.4.3
2.2.4.4
2.2.4.5
2.2.4.6
2.2.4.7
2.2.5
2.2.5.1
2.2.5.2
2.3
2.3.1
2.3.2
2.3.2.1
2.3.2.2
2.3.2.3
2.3.2.4
2.3.3
2.3.4
2.3.4.1
2.3.4.2
2.4
2.5
2.5.1


Catalytic Enantioselective a-Hydrazination of Carbonyl Compounds
using Azodicarboxylates 46
Stereoselective a-Hydrazination of Chiral a,b-Unsaturated
Carboxylates using Azodicarboxylates 50
Disconnection 2: Synthesis from Chiral Nonracemic a-Amino
Acids 52
Schestakow Rearrangement of Hydantoic Acids Prepared from
a-Amino Acids 52
Reduction of N-Nitroso-a-Amino Esters 52
Amination of a-Amino Acids by Hydroxylamine Derivatives 52
Amination of a-Amino Acids by Oxaziridines 53
Disconnections 3, 4, and 5: Syntheses from Hydrazones or
a-Diazoesters 55
Catalytic Enantioselective Hydrogenation of Hydrazones 56
Stereoselective and Catalytic Enantioselective
Strecker Reaction 56
Stereoselective Addition of Organometallic Reagents to
Hydrazones 57
Stereoselective or Catalytic Enantioselective Mannich-Type
Reaction with Hydrazones 58
Enantioselective Friedel–Crafts Alkylations with Hydrazones 59
Diastereoselective Zinc-Mediated Carbon Radical Addition
to Hydrazones 59
Catalytic Enantioselective Reaction of a-Diazoesters with Aldehydes
and Subsequent Stereoselective Reduction 59
Piperazic Acid and Derivatives by Cycloaddition Reactions 61
Diels–Alder Cycloaddition 61
1,3-Dipolar Cycloaddition 62
Chemistry 63
Cleavage of the N–N Bond 63

Reactivity of the Hydrazino Function 67
Reaction of Unprotected a-Hydrazino Acid Derivatives with
Acylating Reagents 67
Reaction of N1-Substituted a-Hydrazino Acid Derivatives with
Acylating Reagents 69
Reaction of N2-Protected a-Hydrazino Acid Derivatives with
Acylating Reagents 69
Reaction with Aldehydes and Ketones 69
Reactivity of the Carboxyl Function 73
Synthesis of Heterocycles 74
Cyclization Leading to Piperazic Acid Derivatives 74
Other Heterocycles 75
Conclusions 78
Experimental Procedures 79
(S)-2-hydrazinosuccinic Acid Monohydrate 79

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VII


VIII

Contents

2.5.2
2.5.3
2.5.4
2.5.5
2.5.6

2.5.7
2.5.8

3

3.1
3.2
3.2.1
3.2.2
3.2.2.1
3.2.2.2
3.2.2.2.1
3.2.2.2.2
3.2.2.2.3
3.3
3.3.1
3.3.1.1
3.3.1.2
3.3.1.3
3.3.1.4
3.3.1.5
3.3.1.6
3.3.1.7
3.3.2
3.3.3
3.3.4
3.3.5
3.3.6
3.3.7
3.3.8

3.3.9

()-(R)-N1,N2-dibenzyloxycarbonyl-2-hydrazino-2-phenyl Propionic
Acid, Methyl Ester 80
(ỵ)-(R)-N1,N2-Bis(benzyloxycarbonyl)-1-hydrazino-2-oxocyclopentane
Carboxylic Acid, Ethyl Ester 81
()-L-N-Aminovaline 81
(ỵ)-L-N-benzyl-N-(tert-butoxycarbonylamino)tryptophan,
Hexylamine Salt 82
(R)-2-(N2-benzoylhydrazino)-2-(4-dimethylaminophenyl)
Acetonitrile 83
tert-Butoxycarbonylamino-(4-dimethylamino-2-methoxy-phenyl)-acetic
Acid Ethyl Ester by Reduction using SmI2 84
(R)-1,2-bis(benzyloxycarbonyl)piperazine-3-carboxylic Acid 84
References 86
Hydroxamic Acids: Chemistry, Bioactivity, and Solutionand Solid-Phase Synthesis 93
Darren Griffith, Marc Devocelle, and Celine J. Marmion
Introduction 93
Chemistry, Bioactivity, and Clinical Utility 93
Chemistry 93
Bioactivity and Clinical Utility 95
Hydroxamic Acids as Siderophores 95
Hydroxamic Acids as Enzyme Inhibitors 97
MMP Inhibitors 98
HDAC Inhibitors 102
PGHS Inhibitors 104
Solution-Phase Synthesis of Hydroxamic Acids 106
Synthesis of Hydroxamic Acids Derived from Carboxylic
Acid Derivatives 106
From Esters 107

From Acid Halides 108
From Anhydrides 109
From [1.3.5]Triazine-Coupled Carboxylic Acids 110
From Carbodiimide-Coupled Carboxylic Acids 111
From Acyloxyphosphonium Ions 111
From Carboxylic Acids Coupled with other Agents 113
Synthesis of Hydroxamic Acids from N-acyloxazolidinones 114
Synthesis of Hydroxamic Acids from gem-Dicyanoepoxides 115
Synthesis of Hydroxamic Acids from Aldehydes 115
Synthesis of Hydroxamic Acids from Nitro Compounds 116
Synthesis of Hydroxamic Acids via a Palladium-Catalyzed
Cascade Reaction 116
Synthesis of N-Formylhydroxylamine (Formohydroxamic Acid) 117
Synthesis of Reverse or Retro-Hydroxamates 117
Synthesis of Acylhydroxamic Acids 120

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Contents

3.4
3.4.1
3.4.1.1
3.4.1.1.1
3.4.1.1.2
3.4.1.2
3.4.1.3
3.4.2
3.4.2.1

3.5
3.6
3.6.1
3.6.2
3.6.3
3.6.4
3.6.5
3.6.6
3.6.7
3.6.8
3.6.9
3.6.10
3.6.11
3.6.12
3.6.13
3.6.14
3.6.15

4
4.1
4.2
4.3
4.4
4.5
4.6
4.7
4.8
4.9
4.10
4.11


Solid-Phase Synthesis of Hydroxamic Acids 121
Acidic Cleavage 122
O-Tethered Hydroxylamine 122
Cleavage with 30–90% TFA 122
Super Acid-Sensitive Linkers 124
N-Tethered Hydroxylamine 126
Other Methods of Solid-Phase Synthesis of Hydroxamic Acids
based on an Acidic Cleavage 126
Nucleophilic Cleavage 128
Other Methods 129
Conclusions 130
Experimental Procedures 130
Synthesis of 3-Pyridinehydroxamic Acid 130
Synthesis of O-benzylbenzohydroxamic Acid 131
Synthesis of N-methylbenzohydroxamic Acid 131
Synthesis of Isobutyrohydroxamic Acid 132
Synthesis of O-benzyl-2-phenylpropionohydroxamic Acid 132
Synthesis of Methyl 3-(2-quinolinylmethoxy)benzeneacetohydroxamic
Acid 133
Synthesis of the Chlamydocin Hydroxamic Acid Analog,
cyclo(L-Asu(NHOH)–Aib-L-Phe–D-Pro) 133
Synthesis of O-benzyl-4-methoxybenzohydroxamic Acid 134
Synthesis of O-benzylbenzohydroxamic acid 134
Synthesis of a 4-chlorophenyl Substituted-a-bromohydroxamic
acid 134
Synthesis of 4-Chlorobenzohydroxamic Acid 135
Synthesis of Acetohydroxamic Acid 135
Synthesis of N-hydroxy Lactam 136
Synthesis of O-tert-butyl-N-formylhydroxylamine 136

Synthesis of Triacetylsalicylhydroxamic Acid 137
References 137
Chemistry of a-Aminoboronic Acids and their Derivatives 145
Valery M. Dembitsky and Morris Srebnik
Introduction 145
Synthesis of a-Aminoboronic Acids 146
Synthesis of a-Amidoboronic Acid Derivatives 146
Asymmetric Synthesis via a-Haloalkylboronic Esters 151
Synthesis of Glycine a-Aminoboronic Acids 154
Synthesis of Proline a-Aminoboronic Acids 155
Synthesis of Alanine a-Aminoboronic Acids 162
Synthesis of Ornithine a-Aminoboronic Acids 164
Synthesis of Arginine a-Aminoboronic Acids 167
Synthesis of Phenethyl Peptide Boronic Acids 170
Synthesis via Zirconocene Species 172

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IX


X

Contents

4.12
4.13
4.14

5

5.1
5.2
5.3
5.3.1
5.3.2
5.3.3
5.3.4
5.4
5.5
5.6
5.6.1
5.6.2
5.6.3
5.6.4
5.6.5
5.6.6
5.7
5.8
5.9
5.9.1
5.9.2
5.9.3
5.9.4
5.9.5

5.9.6

Synthesis and Activity of Amine-Carboxyboranes and
their Derivatives 174
Synthesis of Boron Analogs of Phosphonoacetates 179

Conclusions 183
References 183
Chemistry of Aminophosphonic Acids and Phosphonopeptides 189
Valery P. Kukhar and Vadim D. Romanenko
Introduction 189
Physical/Chemical Properties and Analysis 191
Synthesis of a-Aminophosphonic Acids 193
Amidoalkylation in the ‘‘Carbonyl Compound–Amine–Phosphite’’
Three-Component System 193
Kabachnik–Fields Reaction 195
Direct Hydrophosphonylation of C¼N Bonds 199
Syntheses using C–N and C–C Bond-Forming Reactions 206
Synthesis of b-Aminophosphonates 212
Synthesis of g-Aminophosphonates and Higher Homologs 219
Phosphono- and Phosphinopeptides 227
General Strategies for the Phosphonopeptide Synthesis 229
Peptides Containing P-terminal Aminophosphonate or
Aminophosphinate Moiety 230
Peptides Containing an Aminophosphinic Acid Unit 233
Peptides Containing a Phosphonamide or
Phosphinamide Bond 236
Phosphonodepsipeptides Containing a Phosphonoester Moiety 239
Peptides Containing a Phosphonic or Phosphinic Acid Moiety
in the Side-Chain 240
Remarks on the Practical Utility of Aminophosphonates 240
Conclusions 245
Experimental Procedures 246
Synthesis of N-Protected a-aminophosphinic Acid 10
(R1 ¼ EtOCOCH2, R2 ¼ Me) 246
Synthesis of Phosphonomethylaminocyclopentane-1-carboxylic

Acid (17) 246
General Procedure for Catalytic Asymmetric Hydrophosphonylation.
Synthesis of a-Aminophosphonate 39 (R1 ¼ C5H11, R2 ¼ Ph2CH) 247
General Procedure of the Asymmetric Aminohydroxylation
Reaction: Synthesis of b-Amino-a-hydroxyphosphonates 87 247
Dimethyl (S,S)-(À)3-N,N-bis(a-Methylbenzyl) amino-2oxopropylphosphonate (S,S)-100 and Dimethyl 3-[(S,S)-N,
N-bis(a-methylbenzylamino)-(2R)-hydroxypropylphosphonate
(R,S,S)-101 248
General Procedure for the Preparation of Dialkyl Phenyl(4pyridylcarbonylamino) methyl-phosphonates 126 249

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Contents

5.9.7

Synthesis of 1-[(Benzyloxy) carbonyl] prolyl-N1-{[1,10 -biphenyl-4yl-methyl)(methoxy) phosphoryl] methyl}leucinamide (159a) 249
References 249

6

Chemistry of Silicon-Containing Amino Acids 261
Yingmei Qi and Scott McN. Sieburth
Introduction 261
Stability of Organosilanes 261
Sterics and Electronics 262
Synthesis of Silicon-Containing Amino Acids 263
Synthesis of a-Silyl Amino Acids and Derivatives 263
Synthesis of b-Silylalanine and Derivatives 263

Synthesis of o-Silyl Amino Acids and Derivatives 267
Synthesis of Silyl-Substituted Phenylalanines 269
Synthesis of Amino Acids with Silicon a to Nitrogen 269
Synthesis of Proline Analogs with Silicon in the Ring 269
Reactions of Silicon-Containing Amino Acids 271
Stability of the Si–C Bond 272
Functional Group Protection 272
Functional Group Deprotection 272
Bioactive Peptides Incorporating Silicon-Substituted Amino Acids 272
Use of b-Silylalanine 272
Use of N-Silylalkyl Amino Acids 274
Use of Silaproline 274
Conclusions 275
Experimental Procedures 276
L-b-Trimethylsilylalanine 23 276
(Ỉ)-b-Trimethylsilylalanine 23 276
L-b-Trimethylsilylalanine 23 277
(Ỉ)-p-Trimethylsilylphenylalanine 60 277
L-4-Dimethylsilaproline 100 278
References 278

6.1
6.1.1
6.1.2
6.2
6.2.1
6.2.2
6.2.3
6.2.4
6.2.5

6.2.6
6.3
6.3.1
6.3.2
6.3.3
6.4
6.4.1
6.4.2
6.4.3
6.5
6.6
6.6.1
6.6.2
6.6.3
6.6.4
6.6.5

Part Two Amino Acid Organocatalysis
7
7.1
7.2
7.2.1
7.2.1.1
7.2.1.2
7.2.2
7.2.2.1

281

Catalysis of Reactions by Amino Acids 283

Haibo Xie, Thomas Hayes, and Nicholas Gathergood
Introduction 283
Aldol Reaction 285
Intramolecular Aldol Reaction and Mechanisms 285
Intramolecular Aldol Reaction 285
Mechanisms 287
Intermolecular Aldol Reaction and Mechanisms 289
Intermolecular Aldol Reaction 289

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XI


XII

Contents

7.2.2.2
7.2.3
7.2.3.1
7.2.3.2
7.3
7.3.1
7.3.2
7.3.3
7.3.4
7.4
7.5
7.5.1

7.5.1.1
7.5.2
7.5.2.1
7.5.2.2
7.6
7.6.1
7.6.2
7.7
7.7.1
7.7.2
7.7.3
7.7.4
7.7.5
7.7.6
7.7.7
7.8
7.8.1
7.8.2
7.9
7.10

Mechanisms 292
Carbohydrate Synthesis 294
Carbohydrate Synthesis 294
Synthesis of Amino Sugars 297
Mannich Reaction 298
a-Aminomethylation 298
Direct Mannich Reaction 298
Indirect Mannich Reaction using Ketone Donors 303
anti-Mannich Reactions 303

a-Amination Reaction 306
Michael Reaction 308
Mechanism for Iminium Ion-Catalyzed Michael Reaction 309
Iminium Ion-Catalyzed Intermolecular Michael Reactions 309
Mechanism for the Enamine-Catalyzed Michael Reaction 313
Enamine-Catalyzed Intramolecular Michael Reactions 313
Enamine-Catalyzed Intermolecular Michael Reactions 313
Morita–Baylis–Hillman Reaction and Its Aza-Counterpart 319
Morita–Baylis–Hillman Reactions 319
Aza-Morita–Baylis–Hillman Reactions 320
Miscellaneous Amino Acid-Catalyzed Reactions 321
Diels–Alder Reaction 322
Knoevenagel Condensation 322
Reduction and Oxidation 323
Rosenmund–von Braun Reaction 326
Activation of Epoxides 326
a-Fluorination of Aldehydes and Ketones 327
SN2 Alkylation 328
Sustainability of Amino Acid Catalysis 328
Toxicity and Ecotoxicity of Amino Acid Catalysis 328
Amino Acid Catalysis and Green Chemistry 329
Conclusions and Expectations 330
Typical Procedures for Preferred Catalysis of Reactions
by Amino Acids 330
References 333

Part Three Enzymes
8
8.1
8.2

8.3
8.4

339

Proteases as Powerful Catalysts for Organic Synthesis 341
Andrés Illanes, Fanny Guzmán, and Sonia Barberis
Enzyme Biocatalysis 341
Proteolytic Enzymes: Mechanisms and Characteristics 345
Proteases as Process Catalysts 348
Proteases in Organic Synthesis 350

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Contents

8.5
8.5.1
8.5.2
8.6

Peptide Synthesis 350
Chemical Synthesis of Peptides 351
Enzymatic Synthesis of Peptides 354
Conclusions 360
References 361

9


Semisynthetic Enzymes 379
Usama M. Hegazy and Bengt Mannervik
Usefulness of Semisynthetic Enzymes 379
Natural Protein Biosynthesis 380
Sense Codon Reassignment 381
Missense Suppression 385
Evolving the Orthogonal aaRS/tRNA Pair 387
Nonsense Suppression 390
Mischarging of tRNA by Ribozyme 395
Evolving the Orthogonal Ribosome/mRNA Pair
Frame-Shift Suppression 397
Noncanonical Base Pairs 399
Chemical Ligation 401
Inteins 404
EPL 410
Post-Translational Chemical Modification 411
Examples of Semisynthetic Enzymes 415
Conclusions 419
References 419

9.1
9.2
9.3
9.4
9.5
9.6
9.7
9.8
9.9
9.10

9.11
9.12
9.13
9.14
9.15
9.16

396

Catalysis by Peptide-Based Enzyme Models 433
Giovanna Ghirlanda, Leonard J. Prins, and Paolo Scrimin
10.1
Introduction 433
10.2
Peptide Models of Hydrolytic Enzymes 434
10.2.1
Ester Hydrolysis and Acylation 434
10.2.1.1 Catalytically Active Peptide Foldamers 435
10.2.1.2 Self-Organizing Catalytic Peptide Units 438
10.2.1.3 Multivalent Catalysts 440
10.2.2
Cleavage of the Phosphate Bond 444
10.2.2.1 DNA and DNA Models as Substrates 446
10.2.2.2 RNA and RNA Models as Substrates 450
10.3
Peptide Models of Heme Proteins 456
10.3.1
Heme Proteins 457
10.3.1.1 Early Heme-Peptide Models: Porphyrin as Template 457
10.3.1.2 Bishistidine-Coordinated Models 458

10.3.1.2.1 Water-Soluble Models: Heme Sandwich 458
10.3.1.2.2 Water-Soluble Models: Four-Helix Bundles 460
10.3.1.2.3 Membrane-Soluble Heme-Binding Systems 462
10

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XIII


XIV

Contents

10.3.2
10.4

Diiron Model Proteins: the Due-Ferri Family 464
Conclusions 467
References 467

11

Substrate Recognition 473
Keith Brocklehurst, Sheraz Gul, and Richard W. Pickersgill
Recognition, Specificity, Catalysis, Inhibition, and Linguistics
Serine Proteinases 476
Cysteine Proteinases 480
Glycoside Hydrolases 485
Protein Kinases 488

aaRSs 490
Lipases 492
Conclusions 493
References 494

11.1
11.2
11.3
11.4
11.5
11.6
11.7
11.8

12

12.1
12.2
12.2.1
12.2.2
12.2.3
12.3
12.3.1
12.3.2
12.3.3
12.3.4
12.4
12.4.1
12.4.2
12.4.3

12.4.4
12.4.5
12.5
12.5.1
12.5.2
12.5.3
12.5.4
12.6
12.6.1
12.6.2
12.6.3
12.7

Protein Recognition 505
Robyn E. Mansfield, Arwen J. Cross, Jacqueline M. Matthews,
and Joel P. Mackay
General Introduction 505
Nature of Protein Interfaces 506
General Characteristics of Binding Sites 506
Modularity and Promiscuity in Protein Interactions 507
Hotspots at Interfaces 508
Affinity of Protein Interactions 509
Introduction 509
‘‘Irreversible’’ Interactions 510
Regulatory Interactions 510
Ultra-Weak Interactions 511
Measuring Protein Interactions 512
Introduction 512
Discovering/Establishing Protein Interactions 512
Determining Interaction Stoichiometry 513

Measuring Affinities 514
Modulation of Binding Affinity 515
Coupled Folding and Binding 515
Introduction 515
Characteristics of Intrinsically Unstructured Proteins 516
Advantages of Disorder for Protein Recognition 516
Diversity in Coupled Folding and Binding 518
Regulation of Interactions by PTMs 519
Introduction 519
Types of PTMs 519
A Case Study – Histone Modifications 520
Engineering and Inhibiting Protein–Protein Interactions 521

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473


Contents

12.7.1
12.7.2
12.7.3
12.7.4
12.7.5
12.7.6
12.8

Introduction 521
Engineering Proteins with a Specific Binding Functionality 521

Optimizing Protein Interactions 523
Engineering DNA-Binding Proteins 523
Searching for Small-Molecule Inhibitors of Protein Interactions 524
Flexibility and Allosteric Inhibitors 526
Conclusions 527
References 527

13

Mammalian Peptide Hormones: Biosynthesis and Inhibition 533
Karen Brand and Annette G. Beck-Sickinger
Introduction 533
Mammalian Peptide Hormones 534
Biosynthesis of Peptide Hormones 535
Production and Maturation of Prohormones before Entering the
Secretory Pathway 535
Secretory Pathways 540
Prohormone Cleavage 542
Basic Amino Acid-Specific Members of the Proprotein
Convertase Family 548
Different Biologically Active Peptides from one Precursor 551
Nomenclature at the Cleavage Site 551
Prediction of Cleavage Sites – Discovery of New Bioactive Peptides
Further PTMs 552
Removal of Basic Amino Acids 552
C-Terminal Amidation 553
Acylation 554
Pyroglutamylation 554
N-Terminal Truncation 554
Inhibition of Biosynthesis 555

Readout Systems to Investigate Cleavage by Proteases 555
Rational Design of Inhibitors of the Angiotensin-Converting
Enzyme 557
Proprotein Convertase Inhibitors 561
Endogenous Protein Inhibitors and Derived Inhibitors 562
Peptide Inhibitors 563
Peptide-Derived Inhibitors 563
Are there Conformational Requirements for Substrates? 564
Conclusions 565
References 565

13.1
13.2
13.3
13.3.1
13.3.2
13.3.3
13.3.3.1
13.3.3.2
13.3.3.3
13.3.3.4
13.3.4
13.3.4.1
13.3.4.2
13.3.4.3
13.3.4.4
13.3.4.5
13.4
13.4.1
13.4.2

13.4.3
13.4.3.1
13.4.3.2
13.4.3.3
13.4.3.4
13.5

14
14.1
14.2

Insect Peptide Hormones 575
R. Elwyn Isaac and Neil Audsley
Introduction 575
Structure and Biosynthesis of Insect Peptide Hormones

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576

552

XV


XVI

Contents

14.3

14.4
14.5
14.6
14.7
14.8

Proctolin 578
Sex Peptide 580
A-Type Allatostatins 582
CRF-Related Diuretic Hormones (DH) 584
Insect Peptide Hormones and Insect Control
Conclusions 589
References 590

15

Plant Peptide Signals 597
Javier Narváez-Vásquez, Martha L. Orozco-Cárdenas, and Gregory Pearce
Introduction 597
Defense-Related Peptides 599
Systemin 599
Hydroxyproline-Rich Systemin Glycopeptides 603
Arabidopsis AtPep1-Related Peptides 604
Peptides Involved in Growth and Development 605
CLAVATA3 and the CLE Peptide Family 605
CLAVATA3 (CLV3) 605
CLV3-Related Peptides 607
Rapid Alkalinization Factor Peptides 609
Rotundifolia4 and Devil1 610
C-Terminally Encoded Peptide 1 611

Tyrosine-Sulfated Peptides 611
Phytosulfokine 611
Plant Peptides Containing Sulfated Tyrosine 1 613
Polaris 613
Inflorescence Deficient in Abscission 614
4-kDa Peptide 615
Peptides Involved in Self-Recognition 615
S-Locus Cysteine Rich Peptides 615
Methods in Plant Regulatory Peptide Research 616
Discovery of Systemin 617
Identification of Novel Peptide Signals using the Cell
Alkalinization Assay 618
Isolation of Tyrosine-Sulfated Peptides 621
Use of Peptidomics 622
Fishing Ligands with Bait Receptors 622
Conclusions 623
References 624

15.1
15.2
15.2.1
15.2.2
15.2.3
15.3
15.3.1
15.3.1.1
15.3.1.2
15.3.2
15.3.3
15.3.4

15.3.5
15.3.5.1
15.3.5.2
15.3.6
15.3.7
15.3.7.1
15.4
15.4.1
15.5
15.5.1
15.5.2
15.5.3
15.5.4
15.5.5
15.6

16
16.1
16.2
16.3

Nonribosomal Peptide Synthesis
Sean Doyle
Introduction 631
NRPs 632
NRP Synthetase Domains 635

631

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586


Contents

16.3.1
16.3.2
16.3.3
16.4
16.4.1
16.5
16.5.1
16.5.2
16.5.2.1
16.5.2.2
16.5.2.3
16.6
16.7

Adenylation Domains 635
Thiolation Domains 638
Condensation Domains 638
PPTases 639
40 PPTase Activity Determination 640
Experimental Strategies for NRPS Investigations 642
Degenerate PCR 645
Determination of Adenylation Domain Specificity 647
Protein MS 647
Identification of NRP Synthetase Adenylation

Domain Specificity (Strategy I) 648
Identification of NRP Synthetase Adenylation
Domain Specificity (Strategy II) 649
Non-NRPS 649
Conclusions 650
References 650
Index

657

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XVII


XIX

List of Contributors
Neil Audsley
The Food and Environment
Research Agency
Sand Hutton
York YO41 1LZ
UK

Keith Brocklehurst
Queen Mary, University of London
School of Biological and Chemical
Sciences
Fogg Building, Mile End Road

London E1 4NS
UK

Sonia Barberis
Universidad Nacional de San Luis,
Chacabuco y Pedernera
Faculty of Chemistry, Biochemistry and
Pharmacy
San Luis
Argentina
Annette G. Beck-Sickinger
Leipzig University
Institute of Biochemistry
Brüderstraße 34
04103 Leipzig
Germany
Karen Brand
Leipzig University
Institute of Biochemistry
Brüderstraße 34
04103 Leipzig
Germany

Arwen J. Cross
University of Sydney
School of Molecular and Microbial
Biosciences
G08 Biochemistry Building
NSW 2006
Sydney

Australia
Valery M. Dembitsky
The Hebrew University of Jerusalem
School of Pharmacy
Department of Medicinal Chemistry
and Natural Products
PO Box 12065
Jerusalem 91120
Israel

Amino Acids, Peptides and Proteins in Organic Chemistry.
Vol.2 – Modified Amino Acids, Organocatalysis and Enzymes. Edited by Andrew B. Hughes
Copyright Ó 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
ISBN: 978-3-527-32098-1

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XX

List of Contributors

Marc Devocelle
Royal College of Surgeons in Ireland
Centre for Synthesis & Chemical
Biology
Department of Pharmaceutical &
Medicinal Chemistry
123 St. Stephens Green
Dublin 2

Ireland
Sean Doyle
National University of Ireland
Maynooth
Department of Biology
Maynooth, Co. Kildare
Ireland
Nicholas Gathergood
Dublin City University
School of Chemical Sciences and
National Institute for Cellular
Biotechnology
Glasnevin, Dublin 9
Ireland
Giovanna Ghirlanda
Arizona State University
Department of Chemistry and
Biochemistry
Tempe, AZ 85287-1604
USA
Darren Griffith
Royal College of Surgeons in Ireland
Centre for Synthesis & Chemical
Biology
Department of Pharmaceutical &
Medicinal Chemistry
123 St. Stephens Green
Dublin 2
Ireland


Sheraz Gul
European ScreeningPort GmbH
Schnackenburgallee 114
22525 Hamburg
Germany
Fanny Guzmán
Pontificia Universidad Católica de
Valparso
Institute of Biology
Avenida Brasil 2950
Valparso
Chile
R. Elwyn Isaac
University of Leeds
Institute of Integrative and Comparative
Biology
Faculty of Biological Sciences
Leeds LS2 9JT
UK
Thomas Hayes
Dublin City University
School of Chemical Sciences and
National Institute for Cellular
Biotechnology
Glasnevin
Dublin 9
Ireland
Usama M. Hegazy
Uppsala University
Biomedical Center

Department of Biochemistry and
Organic Chemistry
Box 576
751 23 Uppsala
Sweden

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List of Contributors

Andrés Illanes
Pontificia Universidad Católica de
Valparso
School of Biochemical Engineering
Avenida Brasil 2147
Valparaíso
Chile

Bengt Mannervik
Uppsala University
Biomedical Center
Department of Biochemistry and
Organic Chemistry
Box 576
751 23 Uppsala
Sweden

Uli Kazmaier
Universität des Saarlandes

Institut für Organische Chemie
Im Stadtwald
66123 Saarbrücken
Germany
Valery P. Kukhar
National Academy of Sciences of
Ukraine
Institute of Bioorganic Chemistry and
Petrochemistry
Murmanskaya Street
Kiev 94
Ukraine
Joel P. Mackay
University of Sydney
School of Molecular and Microbial
Biosciences
G08 Biochemistry Building
NSW 2006
Sydney
Australia
Robyn E. Mansfield
University of Sydney
School of Molecular and Microbial
Biosciences
G08 Biochemistry Building
NSW 2006
Sydney
Australia

Celine J. Marmion

Royal College of Surgeons in Ireland
Centre for Synthesis and Chemical
Biology
Department of Pharmaceutical and
Medicinal Chemistry
123 St. Stephens Green
Dublin 2
Ireland
Jacqueline M. Matthews
University of Sydney
School of Molecular and Microbial
Biosciences
G08 Biochemistry Building
NSW 2006
Sydney
Australia
Javier Narváez-Vásquez
University of California Riverside
Department of Botany and Plant
Sciences
3401 Watkins Dr.
Riverside, CA 92521
USA
Martha L. Orozco-Cárdenas
University of California Riverside
Department of Botany and Plant
Sciences
3401 Watkins Dr.
Riverside, CA 92521
USA


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XXI


XXII

List of Contributors

and
University of California Riverside
Plant Transformation Research Center
Riverside, CA 92521
USA
Gregory Pearce
Washington State University
Institute of Biological Chemistry
Pullman, WA 99164
USA
Richard W. Pickersgill
Queen Mary, University of London
School of Biological and Chemical
Sciences
Joseph Priestley Building
Mile End Road
London E1 4NS
UK
Leonard J. Prins
University of Padova

Padova Section
Department of Chemical Sciences and
ITM-CNR
Via Marzolo 1
35131 Padova
Italy
Yingmei Qi
Temple University
Department of Chemistry
1901 N. 13th Street
Philadelphia, PA 19122
USA
Vadim D. Romanenko
National Academy of Sciences of
Ukraine
Institute of Bioorganic Chemistry and
Petrochemistry
Murmanskaya Street
Kiev 94
Ukraine

Paolo Scrimin
University of Padova
Padova Section
Department of Chemical Sciences and
ITM-CNR
Via Marzolo 1
35131 Padova
Italy
Scott McN. Sieburth

Temple University
Department of Chemistry
1901 N. 13th Street
Philadelphia, PA 19122
USA
Morris Srebnik
The Hebrew University of Jerusalem
School of Pharmacy
Department of Medicinal Chemistry
and Natural Products
PO Box 12065
Jerusalem 91120
Israel
Joëlle Vidal
Université de Rennes 1
CNRS UMR 6510, Chimie et
Photonique Moléculaires
Campus de Beaulieu, case 1012
35042 Rennes Cedex
France
Haibo Xie
Dublin City University
School of Chemical Sciences and
National Institute for Cellular
Biotechnology
Glasnevin, Dublin 9
Ireland

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Part One
Synthesis and Chemistry of Modified Amino Acids

Amino Acids, Peptides and Proteins in Organic Chemistry.
Vol.2 – Modified Amino Acids, Organocatalysis and Enzymes. Edited by Andrew B. Hughes
Copyright Ó 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
ISBN: 978-3-527-32098-1

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j3

1
Synthesis and Chemistry of a,b-Didehydroamino Acids
Uli Kazmaier

1.1
Introduction

Although a,b-didehydroamino acids (DDAAs), where the term “didehydro-” is used
to indicate the lack two hydrogen atoms, do not belong to the group of proteinogenic
amino acids, they are commonly found in nature as building blocks of didehydropeptides (DDPs), mainly as secondary metabolites of bacteria and fungi or other
lower organisms. Most of these compounds show interesting biological activities,
such as the b-lactam antibiotics of the cephalosporin group [1], the herbicidal
tetrapeptide tentoxin [2], or the antitumor agent azinomycin A (carzinophilin) [3]
(Figure 1.1).
From a chemical point of view, DDAAs are interesting candidates for the synthesis
of complex amino acids (e.g., via additions to the double bond). Therefore, it is not

surprising that the research on this important class of amino acids has been reviewed
frequently (e.g., by Schmidt [4], Chamberlin [5], and K€
onig et al. [6]). This chapter
gives an overview of the different protocols for the synthesis of DDAAs and their
typical reaction behavior.

1.2
Synthesis of DDAAs
1.2.1
DDAAs via Eliminations
1.2.1.1 DDAAs via b-Elimination
1.2.1.1.1 From b-Hydroxy Amino Acids The elimination of water from the corresponding b-hydroxy amino acids is a straightforward approach towards DDAAs,
especially if the required hydroxy acids are readily available such as serine and
threonine. On elimination didehydroalanine (DAla) and didehydroaminobutenoate

Amino Acids, Peptides and Proteins in Organic Chemistry.
Vol.2 – Modified Amino Acids, Organocatalysis and Enzymes. Edited by Andrew B. Hughes
Copyright Ó 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
ISBN: 978-3-527-32098-1

www.pdfgrip.com


j 1 Synthesis and Chemistry of a,b-Didehydroamino Acids

4

Ph

O

H
N

R

S

O

NH N

N

O

O

O
O

O

O

O

H
N

N

H

O
AcO

N HN

COOH

O

Cephalosporines

O

O

HO
Azinomycin A

Tentoxin

Figure 1.1 Naturally occurring DDPs.

H
N

O

O


CONH2

N
H

H
N

HN
N
H

S

O

O

O

H
N

N
N

NH
NH


NH

O

HO
N

O

N
H

H
N

N

N
H

N

HN

R

O O

O


O
O
O

O

N

N

S

S

O

O

CONH2

O

N
H

HN
N

O


NH

NH

HN
O

O

N

OH
HN

N
H

O

OH

N
S

OH

N

O


S

HN
O

Geninthiocin
R=H
Berninamycin A R = CH3

OH OH
Thiostrepton

Figure 1.2 Naturally occurring didehydroalanine- and
didehydroaminobutenoate-containing peptides.

(DAbu) are formed, two DDAAs also found widely in nature, such as in geninthiocin [7], berninamycin A [8], or thiostrepton (Figure 1.2) [9].
A wide range of reagents can be used for the activation of the OH group and
elimination occurs in the presence of a suitable base. Useful combinations are oxalyl
chloride [10], (diethylamino)sulfur trifluoride [11], dichloroacetyl chloride [12], tosyl
chloride [13], and pyridine or NEt3. PPh3/diethyl azodicarboxylate [14] and carbodiimides in the presence of CuCl [15] can be used as well, and in general the
thermodynamically more stable (Z) isomer is formed preferentially [16]. With respect
to an application of this approach towards the synthesis of natural products, a
stereoselective protocol is required, providing either the (E)- or (Z)-DDAA. Sai et al.
reported a high selectivity for the (E)-DAbu from threonine by using 1-ethyl-3-(3dimethylaminopropyl)carbodiimide (EDC) in the presence of CuCl2, while the (Z)
isomer was obtained from allo-threonine (Scheme 1.1) [17]. Short reaction times

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