Note
Cite This: J. Org. Chem. 2018, 83, 7528−7533
pubs.acs.org/joc
De Novo Design To Synthesize Lanthipeptides Involving Cascade
Cysteine Reactions: SapB Synthesis as an Example
Huai Chen,† Yuan Zhang,† Qian-Qian Li,† Yu-Fen Zhao,† Yong-Xiang Chen,† and Yan-Mei Li*,†,‡
†
Key Lab of Bioorganic Phosphorus Chemistry & Chemical Biology, Department of Chemistry, Tsinghua University, Beijing
100084, P.R. China
‡
Beijing Institute for Brain Disorders, Beijing 100069, P.R. China
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S Supporting Information
*
ABSTRACT: Lanthipeptides are a family of ribosomally synthesized
peptides that have crucial biological functions. However, due to their
complicated structures, the total synthesis of lanthipeptides is
challenging. Here, a novel strategy to construct lanthipeptides is
described, which involves cascade reactions of cysteine, including
Cys disalkylation elimination, Michael reaction, and native chemical
ligation. We utilized this strategy to synthesize lanthipeptide SapB as
an example. This methodology has the potential to obtain
lanthipeptides and their analogues for biological research and drug
discovery.
R
ibosomally synthesized and post-translationally modified
peptides (RiPPs), which have a variety of structures and
functions, have attracted increasing attention for biological
research and drug discovery in recent years.1 Lanthipeptides,
which are widely produced by various bacteria, are one of the
largest and best-studied families of RiPPs.2 Lanthionine (Lan)
and/or 3-methyllanthionine (MeLan) motifs are the characteristic structural features in lanthipeptides (Figure 1a).3 These
thioether-containing cross-links result in the complicated
polycyclic topologies of lanthipeptides. These thioether bridges
are formed by the Michael-type addition reaction between
cysteine (Cys) and dehydroalanine (Dha) or dehydrobutyrine
(Dhb), which is usually generated from dehydration of serine
(Ser) or threonine (Thr) with the assistance of enzymes in the
producing organisms (Figure 1a).2b,4 Many lanthipeptides have
antimicrobial ability against various Gram-positive bacteria,
even some antibiotic-resistant bacteria.2b,5 In addition, some
lanthipeptides have some other biofunctions.2b,6 For example,
duramycin and cinnamycin can inhibit phospholipase A2 in
eukaryotic organisms to maintain normal lipid metabolism;7
labyrinthopeptin A1 manifests favorable activities against HIV
and HSV;8 SapB and SapT may function as biosurfactants to
promote the growth of aerial hyphae.9 Therefore, obtaining
lanthipeptides for biological and medicinal research is a
promising field.
Nevertheless, it is difficult to isolate lanthipeptides from
natural sources for in-depth study and therapeutic use due to
the small amount of production.10 Therefore, various methods
have been developed, both biologically and chemically, to
produce lanthipeptides and their analogues.3,11 Among these
methods, total chemical synthesis provides a powerful
approach to bypass the biosynthetic system, expanding a
more extensive chemical space for biological research and drug
development. However, because of the complicated structures,
total synthesis of lanthipeptides remains a formidable
challenge. To tackle the challenge, several advances have
been reported. Shiba and co-workers reported a pioneering
work for the total synthesis of a natural lanthipeptide, nisin A,
Figure 1. (a) Structures of Dha, Dhb, Lan, and MeLan. Construction
of Lan/MeLan residues in biological systems. (b) Structure of SapB
(1). The green residues are Dha, and the red residues are Lan. The
stereochemistry of the sites with a blue star has not been determined.
Special Issue: Organic and Biocompatible Transformations in
Aqueous Media
© 2018 American Chemical Society
Received: January 29, 2018
Published: June 12, 2018
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DOI: 10.1021/acs.joc.8b00259
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Note
The Journal of Organic Chemistry
in solution phase in 1988 (Scheme 1a).12 However, the
approach was impeded for further development due to its
two Dha residues in its molecule (Figure 1b).9a The relatively
simple topology and post-translational modifications of SapB
make it a good model to verify the viability of our strategy.
Based on our design, SapB was divided into two segments:
SapB(1−9) and SapB(10−21). During Fmoc-SPPS of the two
segments, we used four types of Cys building blocks with
orthogonal protecting groups. The other amino acid building
blocks were all commonly used in Fmoc-SPPS. The special
Dha residues were formed from Cys in situ through the
elimination reaction under bisalkylating conditions either on
the solid support or in the solution phase.20
The synthesis of SapB(10−21) began with the loading of
Fmoc-Asn(Trt)-OH onto 2-chlorotrityl resin, followed by
standard coupling of the next 11 residues sequentially, with a
Fmoc-Cys(Trt)-OH at site 20, a Fmoc-Cys(Acm)-OH at site
16, a Fmoc-Cys(StBu)-OH at site 13, and a Boc-Thz-OH at
site 10 (Figure 2a). These cysteine residues could be
Scheme 1. Strategies for the Total Synthesis of
Lanthipeptides
tedious procedure and low yield. Chemical synthesis of
lanthipeptides on solid resins has also been explored. Tabor
and co-workers reported several approaches to achieve
orthogonally protected Lan/MeLan derivatives, which could
be used to synthesize lanthipeptides fragments via 9fluorenylmethoxycarbonyl solid-phase peptide synthesis
(Fmoc-SPPS).13 Vederas’ group and van der Donk’s group
also utilized the orthogonally protected Lan/MeLan to
construct several full-length lanthipeptides, such as lacticin S,
lacticin 481, epilancin 15X, and their analogues (Scheme
1b).10,14 However, the overall yields were not high enough,
and the synthesis of orthogonally protected building blocks
was still challenging. The biomimetic method, which forms the
Lan/MeLan by intermolecular Michael addition reaction, has
also been approached both in solid phase and in solution.15
When there are multiple Cys and Dha/Dhb in the peptide
sequence, it is difficult to construct the correct polycyclic
structure of the target lanthipeptide by this method.15c To
overcome these obstacles, it is necessary to develop a facile and
effective method to achieve the synthesis.
We found that the generation of Dha16 and Lan11a could be
formed chemically in peptides through cysteine-based
reactions. Thus, we de novo designed a novel strategy to
synthesize full-length lanthipeptides utilizing several commercially available orthogonally protected Cys building blocks.
Through cascade cysteine-based reactions, including Cys
elimination to Dha in situ,16 biomimetic Michael addition
reaction,15,17 and native chemical ligation (NCL),18 multiple
Lan bridge cycles and Dha residues in the peptides can be
achieved (Scheme 1c). With the ability of orthogonal
deprotections, we can accurately control the reaction sites of
Cys and Dha to form the desired structures. In addition, all of
the amino acid building blocks are commercially available.
Therefore, this strategy has the potential to construct
complicated polycyclic topologies of lanthipeptides and their
analogues through de novo design.
The model lanthipeptide we chose was SapB, which was
discovered from Streptomyces coelicolor in 1991.19 Although the
exact structure of SapB has not been reported,4d this 21 amino
acid peptide was determined to have two Lan bridge rings and
Figure 2. (a) Synthesis of SapB(10−21). (b) Synthesis of SapB(1−9)
peptide hydrazide. (c) HPLC traces of 3 and 4 at 215 and 254 nm.
(d) CD spectra of 3 and 4. (e) HPLC trace (215 nm) and ESI-MS
data of 5. (f) HPLC trace (215 nm) and ESI-MS data of 7.
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The Journal of Organic Chemistry
environment.4c,15d,21 The former diastereoisomer in HPLC
was selected as 9 for the following reactions. The last two Cys
residues (Cys6 and Cys16) with Acm groups were deprotected
with CH3COOAg in a solution of H2O/CH3COOH (v/v =
1:1) at 37 °C for 6 h followed by adding DTT.24 The obtained
peptide 10 was treated with DBAA to convert the two free Cys
to Dha (Figure 3a). The reaction was monitored with
deprotected orthogonally when we needed to achieve different
purposes. With the full sequence assembled on the solid resin,
we first chose the residue Cys13 with a StBu group as the
precursor of Dha, as the StBu could be easily removed by
reducing reagent on resin. 2 was treated with dithiothreitol
(DTT) and N,N-diisopropylethylamine (DIPEA) in DMF to
afford the free thiol. Afterward, 2,5-dibromoadipamide
(DBAA) was used to convert the free thiol into the desired
Dha.16 Followed by the standard global deprotection and resin
cleavage using the TFA cocktail, the Dha13 containing peptide
3 was obtained. The crude peptide was purified with
semipreparative RP-HPLC to give the purified 3. Because
the Trt group was removed by the TFA cocktail, the second
free Cys20 was released. Thus, there was a Cys and a Dha
existing simultaneously in 3, and Michael addition reaction
could be induced. We dissolved 3 in alkaline aqueous buffer A
(50 mM Na2HPO4, pH 8.5) at room temperature and shook it
for 1.5 h to induce the Michael reaction to form the first Lan
bridge ring. As the molecular weight of 3 and 4 was the same,
these two molecules could not be identified by MS. We
monitored the reaction with analytical RP-HPLC and observed
that the retention time had a small shift detected with the
absorption at 215 nm, which suggested a new compound was
produced. When detected at 254 nm, the absorption peak
almost disappeared compared to 3, which might be due to the
consumption of Dha (Figure 2c). We also affirmed that there
was no free thiol in the new compound by testing it with
Ellman’s reagent (Figure S7). The circular dichroism (CD)
spectra showed that the new compound had a more helical
structure than peptide 3 (Figure 2d). All together, these results
indicated that 3 was fully converted, undergoing a Michael
reaction between Cys20 and Dha13 to form a cyclic peptide 4.
We further analyzed 4 with a C18 analytical column and
observed two diastereoisomers at a ratio about 6:1 (Figure S8).
The stereoselectivity might due to the generation of an
endocyclic enolate with an adjacent chiral center.4c,21 We
treated the major product with MeONH2·HCl and tris(2carboxyethyl)phosphine hydrochloride (TCEP·HCl) at pH
4.0, and the Thz in 4 was deprotected to release the third
Cys10 (Figure 2e). This newly generated N-terminal free Cys
could be used for the following NCL reaction.
Separately, SapB(1−9) peptide hydrazide22 was prepared
through Fmoc-SPPS with a Fmoc-Cys(StBu)-OH at site 3 and
a Fmoc-Cys(Acm)-OH at site 6. The StBu group on 6 was
removed and then converted to Dha3 by the above method,
followed by treatment with a TFA cocktail and purification
with RP-HPLC to give purified 7 (Figure 2b,f).
With the two requisite peptide segments in hand, we
adopted the method reported by Liu23 to approach the NCL.
However, we found that several byproducts appeared under
the conditions. By identifying them with ESI-MS, we
considered that these byproducts were derived from the
Michael addition of 4-mercaptophenylacetic acid (MPAA)
with 8 and N-terminal peptide MPAA thioester (Figure S9).
To reduce the side reactions, we adjusted the pH of the
reaction system from 6.5 to 5.5. Favorably, with the conversion
maintained, the side reactions were significantly inhibited.
After NCL, the second Lan bridge (Dha3 and Cys10) was
formed by dissolving 8 in the alkaline aqueous buffer A and
shaking it for 2 h. We observed two equal peaks in the HPLC
trace, which indicated that the newly generated chiral center
was racemized (Figure S10). The difference between two
Michael addition steps might be due to the different chemical
Figure 3. (a) NCL of peptides 5 and 7 and following reactions. (b)
HPLC trace (215 nm) and ESI-MS data of 1 (SapB).
analytical HPLC and ESI-MS (Figure 3b). After reaction and
purification, the full-length lanthipeptide 1 (SapB) was
obtained, with a yield of 9.8% based on peptide 3 over six
steps.
In summary, we de novo designed a strategy based on
cascade reactions of Cys residues to synthesize full-length
multicyclic lanthipeptides. The multiple Cys residues could be
deprotected orthogonally when we need to achieve desired
reactions. We have demonstrated a facile synthesis of SapB by
this strategy. To the best of our knowledge, this is the first
example to synthesize SapB. This strategy has several
advantages: (1) all the amino acid building blocks needed to
synthesize lanthipeptides are commercially available; (2) with
the controllable orthogonal deprotections, it has the potential
to construct complicated polycyclic topologies that extensively
exist in lanthipeptides by de novo design; (3) given the
reasonable yield of each step, it can be used to obtain a
milligram scale of lanthipeptides. However, the major flaw of
this strategy is the stereochemistry of the Michael addition. It
is known that addition reactions to Dha in peptides and
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The Journal of Organic Chemistry
proteins will lead to epimeric mixtures.16,20a,25 The diastereomeric ratio may be affected by local chemical environment, but
how to control the stereoselectivity needs to be further
explored. In spite of this, the strategy provides a potential and
promising methodology to obtain lanthipeptides and their
analogues for biological research and drug development.
■
[M + 2H]2+ 641.7, found 641.3. MS (ESI+) calcd for 4:
C49H80N14O20S3 [M + H]+ 1282.4, found 1282.8; [M + 2H]2+
641.7, found 641.4. MS (ESI+) calcd for 5: C48H80N14O20S3 [M +
H]+ 1270.4, found 1269.8; [M + 2H]2+ 635.7, found 635.3.
Synthesis of Peptide 7. 2-Chlorotrityl resin (0.80 g, 0.911
mmol/g) was washed with DCM × 3 and DMF × 3 and then
incubated with 5% N2H4·H2O/DMF (v/v) for 30 min twice. After
being washed, 5% CH3OH/DMF (v/v) was used to cap the
unreacted sites on the resin for 10 min. After being washed with
DCM × 3 and DMF × 3, 0.2 mmol Fmoc-Leu-OH was used to
couple the resin with 0.2 mmol HATU, 0.2 mmol HOAt, and 0.4
mmol DIPEA for 60 min. A solution of DMF/Ac2O/pyridine = 3:2:1
was used to cap the unreacted sites for 30 min after coupling. Fmoc
was removed using 20% piperidine solution in DMF. The following
amino acids (4 equiv) were coupled with HATU/HOAt/DIPEA (4
equiv/4 equiv/8 equiv) activation and monitored by Kaiser test
reagent. The coupling reaction was carried out for 45 min, and the
Fmoc deprotection was for 5 min + 10 min. When the last FmocThr(tBu)-OH coupling and Fmoc deprotection were finished, the
resin was incubated with a solution of DTT (5 equiv) and DIPEA (10
equiv) in 5 mL of DMF at 37 °C for 6 h to remove the StBu group.
After being washed with DCM × 3 and DMF × 3, 4 mL of DMF,
K2CO3 (5 equiv), and DBAA (4 equiv) were added to the resin and
incubated at 37 °C overnight. Fifteen milliliters of a TFA cocktail
including TFA/TIPS/H2O = 95:2.5:2.5 was used to cleave the
peptide from the resin. After being precipitated from cold ether, the
precipitate was dissolved in CH3CN/H2O = 1:1 at a concentration of
5 mg/mL, purified with semipreparative RP-HPLC at a gradient of
15−50% B over 30 min, and then lyophilized. Next, 15.25 mg of 7 was
obtained with a yield of 7.6% based on resin loading. 7 was identified
with analytical RP-HPLC at a gradient of 10−50% B over 30 min and
ESI-MS. MS (ESI+) calcd for 7: C42H77N15O11S [M + H]+ 1001.2,
found 1000.7; [M + 2H]2+ 501.1, found 500.9.
Synthesis of SapB (1). Peptide 7 (0.85 mg, 0.85 μmol, 1.55
equiv) was dissolved in 200 μL of buffer B and then placed in a −20
°C ice-salt bath. Ten microliters of NaNO2 aqueous solution (0.85 M,
15.5 equiv) was added into the peptide solution and reacted at −20
°C for 20 min. Next, 2.52 mg of MPAA (14.8 μmol, 27 equiv) was
dissolved in 200 μL of buffer C and then added to the peptide
solution. After 10 s vortex, the pH was adjusted to 5.5, and then 0.70
mg of peptide 5 (0.55 μmol, 1 equiv) was added. The reaction was
carried out on a 37 °C thermostatic metal bath with 300 rpm for 4 h.
One hundred microliters of TCEP·HCl aqueous solution (0.45 M, 50
equiv) was added to the reaction solution and incubated for 5 min.
The reaction solution was monitored using analytical RP-HPLC at a
gradient of 15−50% B over 30 min. With purification using analytical
RP-HPLC at a gradient of 15−50% B over 30 min, 0.51 mg of peptide
8 was obtained after being lyophilized. 8 was dissolved and cyclized in
300 μL of buffer A in a 2 mL Eppendorf tube at 25 °C for 2 h. With
purification using analytical RP-HPLC at a gradient of 15−50% B
over 30 min, the former diastereoisomer in HPLC was selected as 9
and lyophilized. 9 was dissolved in 300 μL of H2O/CH3COOH = 1:1
solution, and 50 equiv of CH3COOAg was added to the solution and
incubated at 37 °C for 6 h. Next, 150 equiv of DTT was used to
quench the reaction by incubating it with the reaction solution for 5
min and then centrifuged for 5 min at 6000 rpm. The supernatant was
purified using analytical RP-HPLC at a gradient of 15−50% B over 30
min. The obtained 10 was dissolved in 200 μL of buffer A, and 20 μL
of DBAA (0.5 M in DMF) was added to the peptide solution. The
resulting solution reacted at 25 °C for 1 h and then 37 °C for 3 h.
With purification using analytical RP-HPLC at a gradient of 15−50%
B over 30 min, 0.17 mg of purified SapB (peptide 1) was obtained
after being lyophilized. The yield was about 9.8% based on the
amount of 3. 8, 9, 10, and 1 were identified with RP-HPLC and ESIMS. MS (ESI+) calcd for 8: C90H153N27O31S4 [M + 2H]2+ 1119.8,
found 1119.6. MS (ESI+) calcd for 9: C90H153N27O31S4 [M + 2H]2+
1119.8, found 1119.8. MS (ESI+) calcd for 10: C84H143N25O29S4 [M
+ 2H] 2+ 1048.7, found 1048.6. MS (ESI+) calcd for 1:
C84H139N25O29S2 [M + 2H]2+ 1014.6, found 1014.6; [M + 3H]3+
676.7, found 676.7.
EXPERIMENTAL SECTION
General. All the chemical reagents used here except DBAA were
purchased from commercial resources and used without further
purification. Amino acids mentioned below were used for peptide
synthesis: Fmoc-Asn(Trt)-OH, Fmoc-Cys(Trt)-OH, Fmoc-Thr(tBu)-OH, Fmoc-Ile-OH, Fmoc-Cys(Acm)-OH, Fmoc-Leu-OH,
Fmoc-Ser(tBu)-OH, Fmoc-Cys(StBu)-OH, Fmoc-Asp(OtBu)-OH,
Fmoc-Gly-OH, Boc-Thz-OH, Fmoc-Ala-OH, Fmoc-Arg(Pdf)-OH.
HPLC-grade CH3CN, CH3OH, and TFA were purchased from
commercial suppliers and used for peptide purification and analysis.
Peptide synthesis was completed manually using a peptide synthesis
bubbler vessel. Peptides were purified using semipreparative reversephase high-performance liquid chromatography (RP-HPLC) at a flow
rate of 10 mL/min after cleaved from resins. The semipreparative
column was a reversed-phase C18 column (YMC, 5 μm, 250 mm ×
20 mm). All the peptides were analyzed using analytical HPLC with a
reversed-phase C8 or C18 column (YMC, 5 μm, 150 mm × 4.6 mm)
at a flow rate of 0.8 mL/min. The two solutions for HPLC are (A)
water with 0.06% TFA and (B) CH3CN/H2O = 4:1 with 0.06% TFA.
Peptides were identified by ESI-MS using a Thermo MSQ Plus singlequadrupole electrospray ionization mass spectrometer. CH3CN/H2O
= 1:1 with 0.06% formic acid was utilized as the solution at a flow rate
of 0.4 mL/min. Reaction buffers used in the procedure: buffer A: 50
mM Na2HPO4, pH 8.5; buffer B: 6 M Gn·HCl, 200 mM Na2HPO4,
pH 3.0; buffer C: 6 M Gn·HCl, 200 mM Na2HPO4, pH 7.4.
Synthesis of Peptide 5. 2-Chlorotrityl resin (1.01 g, 0.911
mmol/g) was swelled in 6 mL of DCM for 0.5 h. Then, 0.25 mmol
Fmoc-Asn(Trt)-OH was dissolved in 5 mL of DCM with 3 drops of
DMF and 220 μL of DIPEA, and this solution was incubated with the
resin in the peptide synthesis bubbler vessel for 2 h. After being
washed with DCM three times, a solution of DCM/CH3OH/DIPEA
= 17:2:1 was used to cap the unreacted sites for 5 min × 3. Fmoc was
removed using 20% piperidine solution in DMF. The following amino
acids (4 equiv) were coupled with HATU/HOAt/DIPEA (4 equiv/4
equiv/8 equiv) activation and monitored by a Kaiser test reagent. The
coupling reaction was carried out for 45 min, and the Fmoc
deprotection was for 5 min + 10 min. When the last Boc-Thz-OH
coupling was finished, the resin was incubated with a solution of DTT
(5 equiv) and DIPEA (10 equiv) in 5 mL of DMF at 37 °C for 6 h to
remove the StBu group. After being washed with DCM × 3 and DMF
× 3, 4 mL of DMF, K2CO3 (5 equiv), and DBAA (4 equiv) were
added to the resin and incubated at 37 °C overnight. Fifteen milliliters
of a TFA cocktail including TFA/TIPS/H2O = 95:2.5:2.5 was used to
deprotect and cleave the peptide from the resin. After being
precipitated from cold ether, the precipitate was dissolved in HFIP
at a concentration of 10 mg/mL, purified with semipreparative RPHPLC at a gradient of 15−50% B over 30 min, and then lyophilized.
Next, 12.02 mg of 3 was obtained with a yield of 3.8% based on resin
loading. Then, 2.00 mg of peptide 3 was dissolved in 1 mL of buffer A
and then divided equally in two 2 mL Eppendorf tubes. The tubes
were placed on at 25 °C thermostatic metal bath and shaken at 300
rpm for 1.5 h. The reaction solution was monitored with an analytical
C8 column RP-HPLC at a gradient of 10−50% B over 30 min, and
full conversion of 3 was observed. The major diastereoisomer was
separated with a C18 analytical column at a gradient of 15−45% B
over 30 min, and then it was treated with MeONH2·HCl (150 equiv)
and TCEP·HCl (40 equiv), followed by adjusting the pH to 4.0, and
then the resulting solution was shaken at 37 °C for another 24 h. After
purification with analytical RP-HPLC at a gradient of 10−50% B over
30 min, 1.51 mg of pure peptide 5 was obtained with the isolated yield
of 75.7%. 3, 4, and 5 were identified with RP-HPLC and ESI-MS. MS
(ESI+) calcd for 3: C49H80N14O20S3 [M + H]+ 1282.4, found 1281.7;
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The Journal of Organic Chemistry
Synthesis of 2,5-Dibromoadipamide. DBAA was synthesized
using the reported method16 with a yield of 43.2%: 1H NMR
(DMSO-d6, 400 MHz) δ = 1.81−2.11 (4H, m, CH2CH2), 4.37 (2H,
m, 2 × CHBr), 7.35 (2H, s), 7.73 (2H, s) (2 × NH2); 13C NMR
(DMSO-d6, 400 MHz) δ = 32.47, 32.58 (2 × CH2), 48.23, 48.51 (2 ×
CHBr), 169.79, 169.85 (2 × CO); MS (ESI+) calcd for DBAA
C6H10Br2N2O2 [M + H]+ 302.9, found 302.9.
CD Analysis of 3 and 4. The peptide solutions were prepared at a
concentration of 0.5 mg/mL in buffer (10 mM NaH2PO4/Na2HPO4,
pH 5.0). The CD spectra were measured in the standard procedure
with a Chirascan Plus CD.
Ellman Reagent Test of 3 and 4. Peptide 3 or 4 (0.10 mg) was
dissolved in 200 μL of buffer (50 mM NaH2PO4/Na2HPO4, pH 5.0).
Ten equivalents of 5,5′-dithiobis(2-nitrobenzoic acid) (Ellman’s
reagent) was added to each peptide solution and then shaken at 25
°C for 30 min. The reactions were monitored using analytical RPHPLC at a gradient of 10−50% B over 30 min and ESI-MS.
■
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ASSOCIATED CONTENT
* Supporting Information
S
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.joc.8b00259.
Copies of the 1H NMR and 13C NMR spectra of DBAA;
analytical HPLC and ESI-MS data of peptides 3, 4, 8, 9,
and 10; analytical HPLC and ESI-MS data of Ellman
reagent test; analytical HPLC and ESI-MS data of the
NCL reaction between 5 and 7; analytical HPLC trace
for the Michael addition reaction of 3 and 8 (PDF)
■
AUTHOR INFORMATION
Corresponding Author
*E-mail: liym@mail.tsinghua.edu.cn.
ORCID
Yong-Xiang Chen: 0000-0003-3518-0139
Yan-Mei Li: 0000-0003-1215-5010
Notes
The authors declare no competing financial interest.
■
■
ACKNOWLEDGMENTS
This work was supported by the National Natural Science
Foundation of China (21332006, 21672126, 81661148047).
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