Document [original]

Natural Product

Reports

rsc.li/npr

Volume 41

Number 6

June 2024

Pages 863–970

ISSN 0265-0568

HIGHLIGHT

Anke Kurreck, Gavin J. Miller et al.

Purine nucleoside antibiotics: recent synthetic advances

harnessing chemistry and biology

Purine nucleoside antibiotics: recent synthetic

advances harnessing chemistry and biology

Jonas Motter,

Caecilie M. M. Benckendorﬀ,

Sarah Westarp,

Peter Sunde-

Brown,

Peter Neubauer,

Anke Kurreck *

and Gavin J. Miller *

Covering: 2019 to 2023

Nucleoside analogues represent one of the most important classes of small molecule pharmaceuticals and

their therapeutic development is successfully established within oncology and for the treatment of viral

infections. However, there are currently no nucleoside analogues in clinical use for the management of

bacterial infections. Despite this, a signiﬁcant number of clinically recognised nucleoside analogues are

known to possess some antibiotic activity, thereby establishing a potential source for new therapeutic

discovery in this area. Furthermore, given the rise in antibiotic resistance, the discovery of new clinical

candidates remains an urgent global priority and natural product-derived nucleoside analogues may also

present a rich source of discovery space for new modalities. This Highlight, covering work published

from 2019 to 2023, presents a current perspective surrounding the synthesis of natural purine

nucleoside antibiotics. By amalgamating recent eﬀorts from synthetic chemistry with advances in

biosynthetic understanding and the use of recombinant enzymes, prospects towards diﬀerent structural

classes of purines are detailed.

1. Introduction

2. Purine C-nucleosides

2.1. Enzymes involved in the biosynthesis of purine C-

nucleosides

2.2. Syndone ribosides as a synthetic entry point to diverse

purine C-nucleosides

3. Purine nucleosides incorporating ribose ring

modications

3.1. Angustmycins A & C

3.2. Aristeromycin

4. Halogenated purine nucleosides

5. Complex purine nucleoside antibiotics

5.1. Peptidyl purine nucleosides

5.2. Aminonucleoside natural products

6. Conclusion and outlook

7. Author contributions

8. Conicts of interest

9. Acknowledgments

10. References

1. Introduction

Nucleoside antibiotics are a diverse subset of microbial natural

products, whose evolution has led to a variety of unusual

structural characteristics which mimic or present motifs related

to the component nucleosides and nucleotides of nucleic

acids.

1–3

Traditionally, structural analogues of nucleosides and

nucleotides have been investigated for anticancer and/or anti-

viral therapeutic potential.

4–10

However, an oen-overlooked

facet of these molecules and related microbial natural prod-

ucts is their antibacterial activity. There has been a recent

increase in interest in nucleoside analogues as a source for

novel antibacterial exploration,

11,12

compounded by the rise of

antibiotic resistance within standard intervention therapies.

Depending on their constituent heterocyclic base, nucleo-

side antibiotics can be divided into pyrimidine and purine

classes and the structures of the canonical adenine- and

guanine-containing systems are highlighted in Fig. 1 (blue box).

These nucleosides consist of a sugar core (D-ribose) linked to the

heterocyclic nucleobase via ab-N-glycosidic bond. When

combined with either adenine (6-aminopurine) or guanine (2-

amino-6-hydroxypurine) the structures comprise adenosine (8)

and guanosine (9), respectively. Modications to these scaﬀolds

present the basis for structurally diverse nucleoside antibiotics,

with examples incorporating C-glycosides (b-C-glycosidic

Chair of Bioprocess Engineering, Institute of Biotechnology, Faculty III Process

Sciences, Technische Universit¨

at Berlin, Ackerstraße 76, D-13355, Berlin, Germany

School of Chemical and Physical Sciences and Centre for Glycoscience, Keele

University, Keele, Staﬀordshire, ST5 5BG, UK. E-mail: g.j.miller@keele.ac.uk

BioNukleo GmbH, Ackerstraße 76, 13355 Berlin, Germany. E-mail: anke.kurreck@

bionukleo.com

Cite this: Nat. Prod. Rep.,2024,41,873

Received 12th October 2023

DOI: 10.1039/d3np00051f

rsc.li/npr

This journal is © The Royal Society of Chemistry 2024 Nat. Prod. Rep.,2024,41,873–884 | 873

Natural Product

Reports

HIGHLIGHT

linkage), ribose ring halogenation and the attachment of

peptides, illustrated further in Fig. 1.

Traditionally, chemical synthesis has served as the primary

tool to access biologically relevant, structurally dened, and

homogenous purine nucleoside antibiotics. However, due to

their structural complexity, such syntheses are oen chal-

lenging and can require multi-step routes to successfully

prepare a given target.

14,15

This can lead to low overall yields of

material. To overcome these issues, considerable progress has

been made in developing innovative synthetic solutions (vide

infra), but also towards elucidating and understanding the

biosynthetic pathways that produce nucleoside antibiotics.

Availability of recombinant enzymes that operate within such

pathways can provide an alternative, biocatalytic approach

towards the synthesis of relevant targets. This has been

reviewed recently for pyrimidine nucleoside antibiotics.

In this Highlight we present and synergise advancesfrom 2019

onwards surrounding the understanding of biosynthetic

pathways/enzymes and related synthetic chemistry platforms that

can deliver purine nucleoside antibiotics. Readers interested in

a comprehensive overview of over a hundred isolated purine

nucleoside antibiotics and their bioactivity are encouraged to visit

Inoso's reports from 1988 and 1991.

1,2

Table 1 presents the anti-

microbial activity of the nucleosides discussed herein.

We have organised these natural products into diﬀerent

sub-classes and consider each individually within

Jonas Motter

Jonas Motter developed

a passion for natural products

while studying polyketide syn-

thases from mushrooms during

his bachelor's at Friedrich

Schiller Universit¨

at in Jena. In

his master's at Freie Universit¨

in Berlin, he focused on the bio-

catalytic synthesis of antiviral

nucleoside analogues. Since

October 2023, he has been

pursuing his PhD at Technische

Universit¨

at Berlin in the Excel-

lence Cluster: Unifying Systems

in Catalysis (UniSysCat) and is also a member of the BIG-NSE/

Einstein Center of Catalysis graduate school. His research inter-

ests primarily revolve around enzymes, such as halogenases, and

their application as environmentally friendly tools for synthesizing

bioactive molecules.

Caecilie M:M:Benckendorff

Caecilie is from Aarhus, Den-

mark. She moved to the UK in

2010 and began her studies at

the University of Aberdeen in

2013, graduating in 2018 with

an MChem. She joined the

Miller group at Keele University

as a PhD student in 2018,

focussing on the chemical

synthesis of uorinated carbo-

cyclic nucleoside analogues.

Completing her studies in 2022,

she stayed on in the Miller group

as a PDRA, where her research is

focussed on the chemical

synthesis of nucleotides.

Sarah Westarp

Sarah Westarp is a doctoral

student at the Chair of Bio-

process Engineering at the

Institute of Biotechnology of the

Technische Universit¨

at Berlin,

Germany, as well being a scien-

tist at the Biotech company

BioNukleo GmbH since 2018.

Her focus being applied bio-

catalysis, she strives to under-

stand and optimize process

performance. The explored

topics cover specic enzyme

activity, reaction equilibria and

downstream processing of enzymatic reactions.

Peter Sunde-Brown

Peter is from the Gold Coast,

Australia. He completed his

B.BiomedSci (Hons) in 2017 at

Griﬃth University's Institute for

Glycomics, where he worked on

phthalate derived natural

product synthesis. Peter

continued his studies at this

facility, completing his PhD

(2018–2022) in the research

group of Assoc. Prof. Todd

Houston. His research focused

on the application of the Mitsu-

nobu reaction on D-fructose,

leading to the development of the potent cyclodehydration reagent

diphenoxytriphenylphosphorane and a practical large-scale prep-

aration of the valuable iminosugar 1-deoxymannojirimycin. Peter

joined the Miller group as a PDRA in April 2023, where his research

is focused on nucleotide synthesis.

874 |Nat. Prod. Rep.,2024,41,873–884 This journal is © The Royal Society of Chemistry 2024

Natural Product Reports Highlight

a framework of forward chemical synthesis and biosynthetic

perspectives. The structural classications covered are high-

lighted in Fig. 1.

2. Purine C-nucleosides

2.1. Enzymes involved in the biosynthesis of purine C-

nucleosides

C-Nucleosides are produced in nature and are characterised

by an unusual carbon–carbon bond between the sugar

backbone and the nucleobase (relative to the native b-N-

glycosidic linkage). As a result, these nucleosides cannot be

degraded by standard salvage pathway enzymes (e.g.,nucleo-

side phosphorylases or hydrolases).

This catabolic stability

makes them an interesting target for drug development. The

biosynthetic pathways for the purine C-nucleosides pyr-

azofurin (1)andformycin(2)haveonlyrecentlybeeneluci-

dated. Liu and colleagues rst identied the gene cluster of 2

in S. kaniharaensis using intensive cosmid library screening in

2019.

From this seminal work the same group, and alongside

the Chen group, independently showed that ForT from S.

candidus and PyrT (also named PyfQ) from S. kaniharaensis are

C-glycoside synthases (Fig. 2a).

31,32

ForT and PyrT were

formally annotated as b-ribofuranosylaminobenzene 5

′

-phos-

phate synthase-like enzymes (b-RFA-P, EC 2.4.2.54), which are

known from the biosynthesis of methanopterin, a cofactor

component in methanogens (anaerobic archaea).

Notably,

ForT is involved in the formycin pathway and PyrT in pyr-

azofurin biosynthesis.

Heterologously expressed ForT or PyrT incubated with

phosphoribosyl pyrophosphate (PRPP) and 4-amino-1H-

pyrazole-3,5-dicarboxylate (ADCP) (10)or4-hydroxy-1H-

pyrazole-3,5-dicarboxylate (HDCP) (13) respectively, led to the

formation of the corresponding carboxypyrazole ribo-C-

nucleotides 11 or 14 (Fig. 2a). Furthermore, the crystal struc-

ture of ForT in complex with PRPP (12) has recently been

resolved to 2.5 Å (PDB ID: 6YQQ), providing the location of

active site residues critical for the recognition of 12 and

catalysis, paving the way for further biocatalytic development

and application of such C-glycoside synthases.

Anke Kurreck

Anke Kurreck is a Postdoctoral

Researcher in the Bioprocess

Engineering group at the Tech-

nische Universit¨

at Berlin, Ger-

many, and has been CEO of the

Biotech company BioNukleo

GmbH since 2014. She holds

a doctoral degree in Microbi-

ology from Martin-Luther-Uni-

versit¨

at of Halle (Germany) and

completed a research internship

at Wageningen University

during her PhD. Subsequently

she worked as a Postdoctoral

Researcher in the Applied Biochemistry Group at the Technische

Universit¨

at Berlin. Her research focusses on applied sciences with

the aim to reduce the environmental burden and to bring (chemo)

enzymatic synthesis routes increasingly into pharmaceutical

application.

Gavin J:Miller

Gavin was born in Newport,

Wales and read Chemistry with

Medicinal Chemistry at UMIST

where he was awarded an

MChem in Medicinal Chemistry.

Continuing his studies at the

University of Manchester, he

undertook a PhD in synthetic

carbohydrate chemistry, investi-

gating multivalency, followed by

a PDRA at St Andrews Univer-

sity. Gavin then worked in

industry, rstly at Ferring Phar-

maceuticals, on the design of

new treatments for primary dysmenorrhea and secondly at Peak-

dale Molecular on the design of an anti-nicotine vaccine. Gavin

returned to academia and the University of Manchester in 2010,

rstly as a PDRA in the Manchester Institute of Biotechnology

developing chemical synthesis approaches to heparan sulfate and

secondly as a xed-term lecturer, in 2014, within the School of

Chemistry. He took up a Lectureship in Organic Chemistry at Keele

in 2016 and was promoted through the ranks to Professor of Bio-

logical Chemistry in 2021. His group's research involves the

chemical and chemoenzymatic synthesis of carbohydrates.

Peter Neubauer

Peter Neubauer is a professor of

Bioprocess Engineering at the

Institute of Biotechnology of the

TU Berlin, Germany. He holds

a doctoral degree in Microbiology

from the University of Greifswald

(Germany) and completed post-

docs at the Royal Institute of

Technology, KTH (Stockholm,

Sweden) and the Universit¨

at of

Halle (Germany). From 2001 to

2008 he was a professor of Bio-

process Engineering at the

University of Oulu (Finland). His

research focusses on bioprocess development with a focus on

recombinant protein production, process analytical technologies,

and laboratory automation, digitalization and FAIR data in the

developmental process for biomolecules. His achievements were

honored in 2023 with the prestigious Thought leader Award from

Agilent.

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2.2. Syndone ribosides as a synthetic entry point to diverse

purine C-nucleosides

In 2020 Van Calenbergh and colleagues disclosed a platform to

access pyrazole C-nucleosides,

including a synthesis of

pyrazofurin and formycin B. The work focused on establishing

access to syndone ribosides (16) (Fig. 2b), which were subjected

to a [3 + 2] dipolar cycloaddition with an alkyne partner, fol-

lowed by extrusion of CO

via a retro-Diels–Alder reaction. This

Fig. 1 Overview of the purine nucleoside natural products considered in this Highlight, along with the chemical structure of the canonical purine

nucleosides, adenosine, and guanosine (blue box, red numbering for ribose ring and purple for heterocycle).

Table 1 The antibiotic activity of nucleosides discussed in this Highlight

Antibiotic Antimicrobial activity Reference

C-Nucleosides Pyrazofurins Broad-spectrum antiviral, high toxicity (mice LD

:10mgkg

−1

)

by blocking de novo pyrimidine synthesis;

human trials for cancer treatment

17 and 18

Formycins Inhibition of X. oryzae at6mgmL

−1

and Mycobacterium

607 at 6 mg mL

−1

; high toxicity (mice LD

: 250–500 mg kg

−1

cumulative toxicity)

Six-carbon framework Angustmycin A and C Inhibitor of GMP synthesis in Gram-positive bacteria, in vitro

inhibition of S. aureus at >10 mg mL

−1

;in vivo activity in

infected mice (CD

angustmycin C: 12.8 mg kg

−1

)

20 and 21

Aristeromycin SAH hydrolase inhibitors, eﬀective against X. oryzae and

P. oryzae in vitro (MIC: 5 mg mL

−1

) and in rice plants in vivo

22 and 23

Halogenated Adechlorin Adenosine deaminase inhibitor, highly eﬀective against

E. faecalis with MIC: 0.005 mgmL

−1

Nucleocidin Anti-trypanosomal activity, high toxicity (mice LD

: 0.2 mg kg

−1

)25

Complex

nucleoside antibiotics

Miharamycin/amipurimycin Inhibitors of P. oryzae (rice blast disease),

MIC amipurimycin: 5 mgmL

−1

26 and 27

A201A Active against Gram-positive bacteria, MIC: 1–8mgmL

−1

LD =lethal dose, values for injections are given; CD =curative/protective dose; GMP =guanosine monophosphate; SAH =S-

adenosylhomocysteine; MIC =minimal inhibitor concentration.

876 |Nat. Prod. Rep.,2024,41,873–884 This journal is © The Royal Society of Chemistry 2024

Natural Product Reports Highlight

aﬀorded N-protected-5-substituted pyrazoles (17) with multiple

orthogonal diversication points. The approach began by

attempting nucleophilic addition to a C1 ribonolactone, fol-

lowed by anomeric dehydrosilylation to deliver the b-syndone

riboside preferentially. However, this process was not eﬃcient,

and the authors instead pursued a direct Lewis acid-mediated

C-glycosylation. This proved eﬀective, albeit delivering a/

bmixtures of syndone ribosides that required separation before

cycloaddition could take place. Several diﬀerent alkyne

coupling partners were then explored to deliver a library of C-

linked pyrazole analogues, demonstrating capability to pursue

future structure–activity-relationship studies around this motif

(Fig. 2c). The utility of the approach was further demonstrated

by completing formal syntheses of formycin B and pyrazofurin.

3. Purine nucleosides incorporating

ribose ring modiﬁcations

The angustmycin and aristeromycin families of purine nucleo-

sides are potent antibiotics, inhibiting the synthesis of

Fig. 2 (a) Biosynthesis of pyrazofurin and formycin cores using the C-glycoside synthases ForT or PyrT. (b) Chemical synthesis of pyrazole C-

nucleosides via syndone ribosides (16); (c) examples of 5-position diversity accessible using this approach; ADCP (10)=4-amino-1H-pyrazole-

3,5-dicarboxylate, HDCP (13)=4-hydroxy-1H-pyrazole-3,5-dicarboxylate, PRPP (12)=phosphoribosyl pyrophosphate, Pi =phosphate, PPi =

pyrophosphate.

Fig. 3 Identiﬁcation of AgmF to access angustmycin A (3) from angustmycin C (21).

This journal is © The Royal Society of Chemistry 2024 Nat. Prod. Rep.,2024,41,873–884 | 877

Highlight Natural Product Reports

guanosine monophosphate (GMP) in Gram-positive bacteria

and S-adenosyl homocysteine (SAH) hydrolases in rice blast

diseases.

20–23

They have been prepared both enzymatically and

chemically; recent discoveries towards both families are

considered sequentially here.

3.1. Angustmycins A & C

Structurally, the angustmycins are analogues of adenosine (8),

but contain an unusual six carbon ketose (b-D-psicofuranose),

incorporating an additional a-hydroxymethyl group at the 1

′

position. Angustmycin A (3)diﬀers from angustmycin C (21)by

an exo-5

′

-alkene in place of the 5

′

-hydroxymethyl substituent

(Fig. 3). Independently, the groups of Kuzuyama and Price &

Chen reported the identication of the angustmycin biosyn-

thetic gene cluster in 2021.

36,37

They identied and showed in

vitro that AgmF (annotated as an SAH hydrolase, EC 3.13.2.1)

was an unusual dehydratase that transformed 21 into 3(Fig. 3).

Additionally, it was shown by Price and Chen that AgmF could

maintain its activity without the addition of NAD

, indicating an

intriguing self-suﬃciency for co-factor recycling. By heat-

treating the enzyme, NAD

was released suggesting that the

cofactor was bound tightly within the active-site pocket. Further

point mutations of the protein conrmed six amino acid resi-

dues to be critical for binding NAD

close to the catalytically

active Lys-185. Finally, the whole biosynthesis pathway was both

reconstructed in E. coli in vivo and with puried enzymes in vitro

to deliver a one-pot synthesis of 3. Whilst 3and 21 have both

been prepared chemically (e.g., starting from D-fructose),

this

recent advancement from a biocatalytic perspective may

surpass such capability, as it oﬀers a direct entry point to

produce these scaﬀolds for wider chemical or enzymatic

diversication.

3.2. Aristeromycin

Aristeromycin (4)diﬀers from canonical systems through the

purine base being bound to a cyclopentane ring (in place of D-

ribofuranose). Such analogues are known substrates and

inhibitors of SAH hydrolases.

However, their biosynthesis and

especially the cyclisation step from cyclic ketose to a poly-

hydroxylated cyclopentane, is intriguing. The Eguchi group

recently demonstrated that the enzyme Ari2 utilises fructose 6-

phosphate as a donor to produce a phosphorylated six carbon

product (Fig. 4a).

Following heterologous expression of Ari2 in

E. coli and purication via aﬃnity chromatography, the struc-

ture of the ve-membered cyclitol phosphate product was

conrmed using NMR; the relative stereochemistry of the

cyclopentane system was supported using nOe analysis.

Furthermore,

P NMR conrmed NAD

as the cofactor for Ari2

and that this could not be replaced by NADP

. Ari2 from S. cit-

ricolor is a myo-inositol-1-phosphate (MIP) synthase ortholog

Fig. 4 (a) Identiﬁcation of Ari2 catalysing the rearrangement of fructose 6-phosphate (22) to a cyclopentane core (23). Red dot tracks C6 through

the skeletal rearrangement. (b) Proposed mechanism for the formation of the cyclopentane core 23.

synthesis of aristeromycin (4), starting from a non-carbohydrate chiral pool material.

878 |Nat. Prod. Rep.,2024,41,873–884 This journal is © The Royal Society of Chemistry 2024

Natural Product Reports Highlight

(EC 5.5.1.4) and recently the same group elucidated the

stereochemistry of this critical reaction. The authors fed 6R-or

6S-(6-

H) glucose (24)toanS. citricolor fermentation culture and

established a diastereoselective proton abstraction from the C6

position within fructose 6-phosphate (as part of the mechanism

to form the new C–C bond), conrming that Ari2 operates in

a MIP synthase fashion (proposed mechanism highlighted in

Fig. 4b).

From a chemical synthesis perspective, successful proce-

dures have been developed from carbohydrate starting mate-

rials such as D-ribose.

More recently an alternative starting

from (−)-2-azabicyclo[2.2.1]hept-5-en-3-one 30 (Vince lactam,

Fig. 4c) was reported by Morken.

When the lactam nitrogen

was protected with an electron withdrawing carbamate pro-

tecting group (such as tert-butyloxycarbamoyl, Boc), subsequent

reduction of the cyclic amide using NaBH

was enabled,

aﬀording a cis-disubstituted cyclopentene 31. Using a Pt(dba)

catalyst, selective diboration of the internal alkene using

(pin)

was accomplished trans to the cis functional groups

aﬀording diborinate 32. The borinate groups were then con-

verted into their respective hydroxyl groups using H

. Overall,

this diastereoselective alkene functionalisation bypassed the

need for hazardous oxidants and the material was converted

into 4, building the adenine base from the pseudo-anomeric

position.

4. Halogenated purine nucleosides

Halogenated nucleoside analogues are commonly encountered

as antiviral and anticancer drugs, exemplied by sofosbuvir and

cladribine. In contrast to these pharmaceuticals, naturally

occurring halogenated nucleosides are rare.

Excitingly, in

2020 Deng and Zhang revealed the rst Fe

-a-ketoglutarate

dependent halogenase to act upon a 2

′

-deoxy nucleotide

framework.

Through re-sequencing the Actinomadura genome,

AdeV was found and the corresponding protein was shown to

catalyse the conversion of 2

′

-deoxyadenosine-5

′

-mono-

phosphate (dAMP, 33) into 2

′

-deoxy-2

′

-Cl-dAMP (34)in vitro

(Fig. 5a). This nding was key, as although the biosynthesis of

adechlorin (35) had been deciphered,

46,47

the responsible

chlorinase was still unknown. Intriguingly, AdeV showed only

11% amino acid similarity to Wel05, a nonheme iron enzyme,

known to chlorinate alkaloids, including 12-epi-hapalindole C.

48,49

Recent crystal structure and point mutation studies have

provided insight into the molecular mechanisms for these

enzymes [PDB IDs: 7W5T (1.76 Å), 7V57 (2.35 Å)] , paving the way

for future enzymatic synthesis of halogenated nucleosides.

50,51

Furthermore, the group of Liu have elucidated the ring-

expanding steps in vitro occurring during coformycin biosyn-

thesis to access the unusual 1,3-diazepine nucleobase

(Fig. 5b).

Fig. 5 (a) Discovery of AdeV to complete stereospeciﬁc2

′

-chlorination of 2

′

-deoxyadenosine monophosphate. (b) Proposed mechanism for the

enzymatic ring expansion during the biosynthesis of coformycin.

dAMP (33)=2

′

-deoxyadenosine monophosphate; Pi =phosphate, PPi =

pyrophosphate, PPPi =triphosphate.

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Advances towards understanding the biosynthesis of nucleo-

cidin (45)(a4

′

-uorinated nucleoside, Fig. 6) is another notable

recent success concerning the origin of halogenated purine

nucleoside antibiotics. First discovered in S. calvus in 1951, this

compound has regained new interest due to its remarkable

biological and structural prole.

The nucleocidin biosynthetic

cluster has been identied and the enzymes catalysing the early

steps of biosynthesis characterised.

More recently, gene

disruption experiments by the Zechel and O'Hagan groups have

independently provided clues towards the enzymatic require-

ments for 4

′

-uorination and 5

′

-sulfamate ester formation.

55–57

Furthermore, an unusual 3

′

-O-b-glucosylated metabolite was

identied by O'Hagan (Fig. 6).

Therefrom, a UDP-Glc depen-

dant glycosyl transferase (NucGT) and a glucosidase (NucGS)

were uncovered and shown to be able to functionalise the 3

′

position of adenosine derivatives. Intriguingly, the glucosidase

could generate 45,suggestingthe3

′

-O-glucosylated substrate

may be the product of the uorination step. However, the

biochemical mechanism for 4

′

-C–F bond formation and the

specic enzyme performing this halogenation step remains

unclear. The chemical synthesis of 45 was established in the

1970s,

and has been reviewed recently elsewhere.

5. Complex purine nucleoside

antibiotics

5.1. Peptidyl purine nucleosides

Thecorestructureofpurinenucleosideantibioticscanbe

signicantly modied by nonribosomal peptide synthases

(NRPS) and polyketide synthases (PKS). The miharamycins

and amipurimycin are exemplar systems, containing a C9

pyranosyl core and a 2-aminopurine. They are produced by S.

miharaensis and S. novoguineensis respectively and recent

reports from the groups of Liu and Tang independently

identied the responsible biosynthetic gene cluster.

60,61

Their

studies indicated that a polyketide synthase catalyses the early

steps of biosynthesis for both miharamycin and amipur-

imycin, towards the high-carbon sugar core, and that within

this MihI acts as bifunctional guanylglucuronic acid synthase

(Fig. 7).

MihI is responsible rst for the anomeric hydrolysis

of GMP (46), followed by use of the released guanine for the

formation of a new C–Nbondvia reaction with the sugar

nucleotide donor, UDP-GlcA. This reaction is particularly

interesting as it incorporates a pyranose in place of ribose.

Furthermore, C–N bond formation in nucleosides is usually

catalyzed by nucleoside phosphorylases or phosphoribosyl

transferases.

Yu and colleagues recently described the rst total synthesis of

amipurimycin (48), completing a 27-step process that adopted

a convergent strategy, coupling the C9 pyranose core to the

respective amino acid and 2-amino purine components.

Impor-

tantly, a comparison of the NMR data from the isolated natural

product to those obtained from previous syntheses highlighted

inconsistencies,

and led to the authors proposing a revised

congurational assignment at C3

′

and C8

′

of this natural product

(Fig. 8a), supported using X-ray crystallography.

63,65

More recently, the chemical synthesis of miharamycin B (6)

and analogues was reported by Wang and Liu.

Starting from

enantiopure (R)-Garner's aldehyde (49) and 3-bromofuran (50),

a 20-step asymmetric de novo approach was designed (Fig. 8b).

Notable from this was coupling of a pyranose-derived interme-

diate (51) with N-Boc-C6-chloropurine using palladium. The

choice of the C6-chloropurine nucleophile proved critical for

obtaining the required N

-regioselectivity; the use of C6-bromo

or iodo purines resulted in oxidative addition of Pd to the

carbon halogen bond, competing with the activation of the

Fig. 6 Discovery of a glucosylated metabolite (44) within nucleocidin biosynthesis.

Fig. 7 Discovery of MihI to complete ribose to glucuronic acid transglycosylation with retained b-anomeric stereochemistry; Pi =phosphate.

880 |Nat. Prod. Rep.,2024,41,873–884 This journal is © The Royal Society of Chemistry 2024

Natural Product Reports Highlight

allylic O-Boc electrophile. The stereocentres within the

remaining sugar framework were installed through reduction of

the C4

′

-ketone (using NaBH

) followed by stereocontrolled

dihydroxylation of the C2

′

–C3

′

alkene using KMnO

to aﬀord 53.

Finally, removal of the C8

′

–C9

′

-acetonide protecting group fur-

nished the desired core structure and enabled completion of

the total synthesis for 6: a series of protecting group manipu-

lations, cyclisation to form the fused ring system, coupling of

arginine and dehalogenation of the nucleobase.

5.2. Aminonucleoside natural products

Another important class of complex purine nucleoside antibi-

otics is the A201 family. Consisting of a 3

′

-deoxy-3

′

-amino

adenosine core, a derivative from this family, A201A (7), was

rst isolated from S. capreolus,

and further steps to elucidate

the biosynthetic pathway in M. thermotolerans (largely con-

cerning the sugar components) have been completed by Ju.

Wang et al. recently reported a total chemical synthesis of

the A201 family, using a convergent strategy reliant on the

Fig. 8 (a) Proposed structural revision to amipurimycin (48) concerning stereochemical assignments at C3

′

and C8

′

of the sugar (highlighted with

red dots). (b) Overview of key steps from the total synthesis of miharamycin B (6).

Fig. 9 Overview of chemical synthesis of A201A (7), highlighting a key diastereoselective b-glycosylation of a galactofuranosyl donor (55); Quin

=quinoline ester.

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coupling of three carbohydrate-based building blocks: a gal-

actofuranose, a 3

′

-amino adenosine and a D-rhamnose.

Underpinning to this synthesis was stereoselective installa-

tion of the 1,2-cis glycosidic linkage to the central galactofur-

anose subunit (Fig. 9). The use of a quinoline ester at the C5-

position proved invaluable, allowing for an elegant b-directed,

Lewis-acid mediated glycosidation of a phenolic acceptor,

proposing coordination of the phenol moiety to the quinoline

nitrogen and aﬀording intermediate 56.Withthedesired

linkage in place, the remaining synthesis of 7was completed

over 9 steps, including: Z-selective formation of the exocyclic

methoxy enol ether, coupling of the 3

′

-amino adenosine

moiety and attachment of the remaining 3,4-di-O-methylated

rhamnose.

6. Conclusion and outlook

Natural products oﬀer a rich source of structural diversity to

discover new nucleoside analogues. Small modications to

such structures can build structure–activity relationships and

probe new druggable space. Classically, following their isola-

tion and structural characterisation, total chemical syntheses

of such molecules would be completed to obtain relevant

quantities of material for further or wider biological investi-

gation. However, as illustrated herein, the length and

complexity of some chemical syntheses (e.g., 27 steps in the

case of 48) demands a substantial amount of resource. It is

here that a synergy of biology (isolation, understanding and

recombinant production of biosynthetic enzymes) and

chemistry (synthesis and structural analysis) may prove

transformative in facilitating the future of purine nucleoside

antibiotic discovery. Indeed, such concepts can be drawn from

examples illustrated herein. An ability to access proteins that

can aﬀectintriguingandchemicallydiﬃcult transformations

–stereospecic2

′

-chlorination (AdeV), furanose to pyranose

transglycosylation (MihI), carbocyclisation (Ari2) –could

rapidly access key core materials for further derivatisation.

Furthermore, the identication of new metabolic enzymes

from within nucleocidin biosynthesisprovidesopportunities

to explore such derivatives and related systems, such as the 7-

deazapurine tubercidin.

70,71

As our ability to make mimetic protein structures prog-

resses, an integration of synthetic biology within classical

purine nucleoside chemical synthesis will be trans-

formative.

Entering a post-antibiotic era predicates a need to

develop new therapeutics. In this context, purine-derived

nucleoside antibiotics are a promising and largely untapped

source of potential. There is a fascinating diversity to this

compound class and several examples have already demon-

strated promising antibacterial activities.

2,73

Realising their

potential will rely on the continued harmony of biocatalysis

with chemical synthesis,

amultidisciplinaryapproach

already delivering across the wider eld of natural product

synthesis.

In recent years, the biocatalytic synthesis of

nucleosides and derivatives has burgeoned, for example by

exploiting the promiscuity and thermostability of nucleoside

phosphorylases,

76–78

and such strategies could be envisaged to

further expand and ease the synthesis of purine nucleoside

antibiotics.

7. Author contributions

Conceptualization: J. M. and P. N.; writing –original dra: J. M.,

C. B., P. S.-B. and G. M. writing –review & editing: J. M., C. B.,

S. W., P. S.-B., P. N., A. K. and G. M.; visualization: J. M. and

G. M.; supervision: P. N., A. K. and G. M.; project administra-

tion: P. N., A. K, and G. M.

8. Conﬂicts of interest

AK is CEO of BioNukleo GmbH. SW is a scientist at BioNukleo

GmbH. PN is a member of the BioNukleo GmbH advisory board.

These aﬃliations constitute no conict of interest with the

topics presented and discussed in this report. The remaining

authors declare no conict of interest.

9. Acknowledgments

G. M. is supported by UK Research and Innovation, who are

thanked for project grant funding (MR/T019522/1 & MR/

W029324/1).

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