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ACKNOWLEDGMENTS
Funding:This work was supported by National Institutes of Health
grant R01 GM081617 (R.Ki.); Israel Science Foundation grant
3055/19 within the Israel Precision Medicine Partnership program
(R.Ki.);, Ernest and Bonnie Beutler Research Program of Excellence
in Genomic Medicine (R.Ki.); European Research Council FP7 ERC
grant 281891 (R.Ki.); Wellcome Trust Sir Henry Wellcome
fellowship 204684/Z/16/Z (M.S.); and D. Dan and Betty Kahn
Foundation’s gift to the University of Michigan, Weizmann Institute,
and Technion–Israel Institute of Technology Collaboration for
Research.Author contributions:M.S., O.S., I.Y., G.K., J.K., G.C.,
V.S., and R.Ki. designed the study. R.Ka. and E.H. curated clinical
data. M.P., G.R., T.W., O.S., and Y.A. collected clinical samples.
G.R., T.W., G.K., and J.K. handled clinical project administration.
M.S., O.S., I.Y., and Y.A. performed whole-genome sequencing. M.S.
and R.Ki. analyzed the data. M.S., I.Y., O.S., B.F., M.P., G.C., V.S.,
and R.Ki. interpreted the results. M.S. and R.Ki. wrote the paper
with comments from all authors.Competing interests:The
authors declare that they have no competing interests.Data and
materials availability:The clinical data that support the findings
of this study are available from Maccabi Healthcare Services,
but restrictions apply to the availability of these data, which were
used under license for the current study and so are not publicly
available. Access to the data is, however, available upon reasonable
request and signing a material transfer agreement with Maccabi
Healthcare Services. Analysis code is available from https://github.com/Technion-Kishony-lab/Antibiotic-treatment-failure ( 35 ). All
urine culture isolate whole-genome sequencing data generated in
this study have been deposited in the Sequence Read Archive
database and are available here:www.ncbi.nlm.nih.gov/sra/
PRJNA786867. Treatment and susceptibility data for the sequenced
isolates are provided in the supplementary materials (data S1).SUPPLEMENTARY MATERIALS
science.org/doi/10.1126/science.abg9868
Materials and Methods
Supplementary Text
Figs. S1 to S17
Tables S1 to S8
References ( 36 – 38 )
MDAR Reproducibility Checklist
Data S110 February 2021; resubmitted 7 September 2021
Accepted 23 December 2021
10.1126/science.abg9868ORGANIC CHEMISTRY
Total synthesis of himastatin
Kyan A. DÕAngelo^1 , Carly K. Schissel^1 , Bradley L. Pentelute1,2,3,4*, Mohammad Movassaghi^1 *The natural product himastatin has an unusual homodimeric structure that presents a substantial synthetic
challenge. We report the concise total synthesis of himastatin from readily accessible precursors, incorporating
a final-stage dimerization strategy that was inspired by a detailed consideration of the compoundÕs biogenesis.
Combining this approach with a modular synthesis enabled expedient access to more than a dozen
designed derivatives of himastatin, including synthetic probes that provide insight into its antibiotic activity.
T
he proliferation of multidrug-resistant
pathogenic bacteria is widely recognized
as a threat to global health ( 1 , 2 ). Natural
products have served as the primary in-
spiration for new antibiotics to treat bac-
terial infections ( 3 ). (–)-Himastatin ( 1 ) is a
macrocyclic peptide with a homodimeric struc-
ture isolated fromStreptomyces himastatinicus
(Fig. 1) that demonstrates antibiotic and anti-
tumor activity ( 4 – 6 ). Although (–)-himastatin’s
( 1 ) mechanism of action is not known, an early
investigation demonstrated that its antibiotic
activity was reduced in the presence of sodium
salts of phospholipids and fatty acids, leading
to speculation that (–)-himastatin ( 1 ) may target
the bacterial membrane ( 7 ). (–)-Himastatin’s
( 1 ) homodimeric structure does not resemble
those of any well-characterized antibiotic class,
including known membrane-disrupting cyclic
peptides. The most distinctive structural fea-
ture of (–)-himastatin ( 1 ) is the central C5–C5′
linkage between cyclotryptophan residues that
is formed in the final biosynthetic step ( 8 )andis
critical for the observed Gram-positive antibiotic
activity ( 9 ). Related monomeric natural products
discovered after (–)-himastatin ( 1 ), including
(–)-NW-G01 (S2), show a substantial enhance-
ment in antibiotic activity upon chemoenzy-
matic dimerization ( 10 ) (fig. S1). Other notable
structural features of (–)-himastatin ( 1 ) include
the alternating sequence ofD- andL-amino acids,
a depsipeptide linkage, and the piperazic acid
residue withg-hydroxylation.
Danishefsky’s landmark synthesis of (–)-
himastatin ( 1 ), which clarified the Castereo-
chemistry of the cyclotryptophan residue,
featured an early-stage Stille coupling to
form the central C5−C5′linkage followed by
bidirectional elaboration of a dimeric cyclo-
tryptophan ( 9 ). Early-stage formation of this
linkage also featured in total syntheses of the
related natural product (–)-chloptosin (S1) by
Yao ( 11 ) and Ley ( 12 ) and their co-workers,
who found that cross-coupling approaches
to achieve a more attractive late-stage dimer-
ization (which would also offer access to hetero-
dimeric derivatives) were not successful ( 12 ).
Motivated by the distinctive structure, established
synthetic challenge, and antibiotic activity, we
became interested in developing a concise total
synthesis of (–)-himastatin ( 1 ) that would offer
rapid access to derivatives for chemical biology
studies.
The key unaddressed challenge of uniting
two complex fragments to form the C5−C5′bond at the center of (–)-himastatin’s( 1 )
dimeric structure encouraged us to consider
the development of a new synthetic methodol-
ogy. To address the Csp2–Csp2linkage present
in (–)-himastatin ( 1 ), we needed to identify a
strategy that stands apart from our group’s
prior approaches based on reductive or pho-
tolytic radical generation and coupling to secure
Csp3–Csp3linkages and Csp3–Csp2linkages be-
tween similar ( 13 ) and dissimilar fragments
( 14 ). We began with a detailed examination of
(–)-himastatin’s( 1 ) biosynthesis from a linear
peptide 4 that is cyclized and then subjected
to oxidative tailoring by three cytochrome P450
enzymes ( 8 ). The final step, catalyzed by HmtS,
forges the central C5–C5′bond by oxidative di-
merization of (+)-himastatin monomer ( 2 ). On
the basis of recent theoretical studies of P450-
catalyzed C–C bond formation, we envisioned
that this enzymatic dimerization may take place
via radical–radical coupling of two cyclotrypto-
phan radicals (fig. S2) ( 15 , 16 ). These radical
species are likely generated in rapid succes-
sion via indoline N–H hydrogen-atom abstrac-
tion at the heme active site, before undergoing
combination in its vicinity ( 16 , 17 ).
We envisioned that a biosynthetically inspired
chemical method for the oxidative dimeriza-
tion of cyclotryptophans could follow the same
radical–radical coupling blueprint. As opposed
to hydrogen atom abstraction, we planned to
generate an analogous open-shell cyclotrypto-
phan species via single-electron oxidation of
the embedded aniline substructure. Consistent
with studies of aniline dimerization via single-
electron oxidation ( 18 – 20 ), we predicted that
the resulting arylamine radical cation would
rapidly dimerize at the most accessible posi-
tion, forming the desired C5–C5′linkage.
Late-stage application of this chemistry to
dimerization of (+)-himastatin monomer ( 2 )
permits a straightforward modular assembly
of linear hexadepsipeptide 5 akin to native
precursor 4 , without the constraints imposed894 25 FEBRUARY 2022•VOL 375 ISSUE 6583 science.orgSCIENCE
(^1) Department of Chemistry, Massachusetts Institute of
Technology, Cambridge, MA 02139, USA.^2 The Koch Institute
for Integrative Cancer Research, MIT, Cambridge, MA 02142,
USA.^3 Broad Institute of MIT and Harvard, Cambridge,
MA 02142, USA.^4 Center for Environmental Health
Sciences, MIT, Cambridge, MA 02139, USA.
*Corresponding author. Email: [email protected] (M.M.);
[email protected] (B.L.P.)
RESEARCH | REPORTS