Engineered biosynthesis of glycosylated derivatives of narbomycin and evaluation of their antibacterial activities

Ah Reum Han, Pramod B. Shinde, Je Won Park, Jaeyong Cho, So Ra Lee, Yeon Hee Ban, Young Ji Yoo, Eun Ji Kim, Eunji Kim, Byung-Gee Kim,Dong Gun Lee, and Yeo Joon Yoon

Table of content

Physico-chemical and spectral data for narbomycin (NB) and its analogues / ∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 2
Table S1 Deoxyoligonucleotide primers used in this study / ∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 3
Table S2NMR data of narbomycin (NB) / ∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 4
Table S3NMR data of D-quinovosyl-narbonolide (DQNVNB)∙∙∙∙ ∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 5
Table S4NMR data of 3-O-demethyl-D-chalcosyl-narbonolide (ODDCNB) / ∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 6
Table S5NMR data of L-rhamnosyl-narbonolide (LRHMNB)∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 7
Table S6NMR data of D-digitoxosyl-narbonolide (DDGTNB)∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 8
Table S7 NMR data of D-boivinosyl-narbonolide (DBVNNB)∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 9
Table S8NMR data of L-oliovosyl-narbonolide (LOLVNB)∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 10
Table S9 NMR data of L-digitoxosyl-narbonolide (LDGTNB)∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 11
References∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙∙ / 12

Physico-chemical and spectral data:

Narbomycin (NB): amorphous white solid; NMR data see Table S2, respectively; ESIMS (+ve mode) m/z 510 [M + NH4]+; HRESIMS m/z 509.3358 [M + H]+ (calcd for C28H47NO7, 509.3353).

D-quinovosyl-narbonolide (DQNVNB): amorphous white solid; NMR data see Table S3, respectively; ESIMS (+ve mode) m/z 516 [M + NH4]+, 335 [M + NH4sugar H2O]+; HRESIMS m/z 516.3170 [M + NH4]+ (calcd for C26H46NO9, 516.3173).

3-O-demethyl-D-chalcosyl-narbomycin (ODDCNB): amorphous white solid; NMR data see Table S4, respectively; ESIMS (+ve mode) m/z 500 [M + NH4]+, 335 [M + NH4sugar H2O]+; HRESIMS m/z500.3221 [M + NH4]+ (calcd for C26H46NO8, 500.3223).

L-rhamnosyl-narbonolide (LRHMNB): amorphous white solid; NMR data see Table S5, respectively; ESIMS (+ve mode) m/z 516 [M + NH4]+, 335 [M + NH4sugar H2O]+; HRESIMS m/z 516.3176 [M + NH4]+ (calcd for C26H46NO9, 516.3173).

D-digitoxosyl-narbonolide (DDGTNB): amorphous white solid; NMR data see Table S6, respectively; ESIMS (+ve mode) m/z 500 [M + NH4]+, 335 [M + NH4sugar H2O]+; HRESIMS m/z500.3225 [M + NH4]+ (calcd for C26H46NO8, 500.3223).

D-boivinosyl-narbonolide (DBVNNB): amorphous white solid; NMR data see Table S7, respectively; ESIMS (+ve mode) m/z 500 [M + NH4]+, 335 [M + NH4sugar H2O]+; HRESIMS m/z500.3228 [M + NH4]+ (calcd for C26H46NO8, 500.3223).

L-olivosyl-narbonolide (LOLVNB): amorphous white solid; NMR data see Table S8, respectively; ESIMS (+ve mode) m/z 500 [M + NH4]+, 335 [M + NH4sugar H2O +]; HRESIMS m/z500.3220 [M + NH4]+ (calcd for C26H46NO8, 500.3223).

L-digitoxosy-narbonolide (LDGTNB): amorphous white solid; NMR data see Table S9, respectively; ESIMS (+ve mode) m/z 500 [M + NH4]+, 335 [M + NH4sugar H2O]+; HRESIMS m/z500.3230 [M+NH4]+ (calcd for C26H46NO8, 500.3223).

1

Table S1 Deoxyoligonucleotide primers used in this study.

Primera / Sequence (5-3) / Restriction enzyme / Source of DNA
(Gene accession number)
desVIII-VII F / TTAATTAAGATATCACCGGCAAGGAAGGACACGACGCC / PacI-EcoRV / Streptomyces venezuelae ATCC 15439
(GenBank AF079762.1)
desVIII-VII R / TCTAGACATATGGCGCAGATACAGGGGTGAGGCCTG / NdeI-XbaI
desI-II F / TTAATTAAACTAGTATCGATGACGGTGGCCCGGAGGG / PacI-SpeI-ClaI / Streptomyces venezuelae ATCC 15439
(GenBank AF079762.1)
desI-II R / TCTAGATGCGGGTCAGCGCAGGAAGCCGCG / XbaI
desIII-IV F / TTAATTAAACTAGTTAACTCGCCACGCCGACCGTT / PacI-SpeI / Streptomyces venezuelae ATCC 15439
(GenBank AF079762.1)
desIII-IV R / TCTAGAGAGCTCCTCGTAGGCGGCCTT / XbaI
desV F / TTAATTAAACTAGTCAGGTCTCCTTCGCGGACGGCCTC / PacI-SpeI / Streptomyces venezuelae ATCC 15439
(GenBank AF079762.1)
desV R / TCTAGAACCTGACTAGGCCTGGTCGACCCG / XbaI
desVI F / TTAATTAAACTAGTCCCCAGGCCTCACCCCTGTATCTG / PacI-SpeI / Streptomyces venezuelae ATCC 15439
(GenBank AF079762.1)
desVI R / TCTAGAGAATTCGCTCAGGCGGGGACGCCGACGAAG / XbaI-EcoRI
oleL F / TTAATTAAACTAGTATCGATATCGCTCCGAGCCCGAAGGGA / PacI-SpeI-ClaI / Streptomyces antibioticus ATCC 11891
(GenBank AF055579.2)
oleL R / TCTAGAGCCGGCCAGTACGAGGGCCTT / XbaI
oleU F / TTAATTAAACTAGTGTACCGCGACAACCGC / PacI-SpeI / S. antibioticus ATCC 11891
(GenBank AF055579.2)
oleU R / TCTAGAGAAGAGGGCCAGGTCGTGCACGC / XbaI
oleV F / TTAATTAAACTAGTATCGATGGGAATCGCGGAAGCG / PacI-SpeI-ClaI / S. antibioticus ATCC 11891
(GenBank AF055579.2)
oleV R / TCTAGAATCAGCTCAGGGCCTGGATGC / XbaI
eryBII F / TTAATTAAACTAGTGGTCGTCGGCATCTGCG / PacI-SpeI / Saccharopolyspora erythraea NRRL 2338
(GenBank U77454.1)
eryBII R / TCTAGACTCGGTTCCTCTTGTGCTCACTGC / XbaI
urdR F / TTAATTAAACTAGTACAGAGATCCAGGACGACGCA / PacI-SpeI / pYJ146 (5)b
urdR R / TCTAGATCAGATACGGACGGCGGAGGT / XbaI
oleW F / TTAATTAAACTAGTAAGGGAACCCCATGCCCTCCC / PacI-SpeI / S. antibioticus ATCC 11891
(GenBank AF055579.2)
oleW R / TCTAGAATCAGCACCAGCGCACCCGCGCCA / XbaI

a F and R represent forward and reverse direction of corresponding primers, respectively.

b pYJ146 is described in Hong et al 2004.

Table S2 NMR data of narbomycin (NB) in CDCl3.

Position / C / H (m, J in Hz)
1 / 169.9
2 / 50.9 / 3.85 (q, 7)
3 / 207.9
4 / 48.9 / 2.85 (m)
5 / 79.0 / 4.18 (brs)
6 / 36.6 / 1.74 (m)
7a
7b / 37.0 / 1.45 (m)
1.18 (m)
8 / 42.2 / 2.76 (m)
9 / 203.8
10 / 128.5 / 6.12 (d, 16)
11 / 147.2 / 6.67 (dd, 15.5, 6)
12 / 38.6 / 2.68 (m)
13 / 79.0 / 4.92 (m)
14 / 22.9 / 1.60 (m)
1.53 (m)
15 / 10.7 / 0.90 (t, 7)
16 / 14.5 / 1.38 (d, 7)
17 / 13.9 / 1.36 (d, 7)
18 / 17.2 / 1.02 (d, 6.5)
19 / 15.7 / 1.12 (d, 7)
20 / 12.8 / 1.09 (d, 7)
1 / 104.6 / 4.33 (d, 7)
2 / 70.3 / 3.31 (dd, 7, 2.5)
3 / 65.8 / 2.94 (m)
4a
4b / 31.2 / 1.60 (m)
1.53 9m)
5 / 69.4 / 3.58 (m)
6 / 21.3 / 1.25 (d, 6)
7 / 40.2 / 2.38 (s)

Table S3NMR data of D-quinovosyl-narbonolide (DQNVNB) in CDCl3.

Position / C / H (m, J in Hz) / COSY / HMBC
1 / 169.7
2 / 50.7 / 3.82 (q, 7.0) / H-16 / C-1, C-3, C-16
3 / 209.0
4 / 48.7 / 2.89 (m, 7.5, 5) / H-5, H-17 / C-3, C-5, C-6, C-17
5 / 78.8 / 4.16 (brs) / H-4
6 / 36.5 / 1.80 (m)
7a
7b / 37.3 / 1.48 (m)
1.19 (m) / C-9, C-5, C-8, C-6
C-9, C-5, C-8, C-6
8 / 41.5 / 2.81 (m, 7) / H-19 / C-9, C-7, C-19
9 / 204.0
10 / 128.8 / 6.05 (d, 16) / H-11 / C-9, C-11, C-13, C-12
11 / 147.5 / 6.65 (dd, 16, 6) / H-10, H-12 / C-9, C-10, C-13, C-12, C-20
12 / 38.6 / 2.72 (m) / H-13, H-20 / C-11, C-10, C-13, C-20
13 / 79.0 / 4.94 (dt, 9.5, 4) / H-12, H-14a, H-14b / C-1, C-11
14a
14b / 23.0 / 1.61 (m)
1.52 (m) / H-13
H-13 / C-15
C-15, C-13
15 / 10.7 / 0.89 (t, 7.5) / C-13, C-14
16 / 14.4 / 1.35 (d, 7.5) / H-2 / C-1, C-2, C-3
17 / 13.5 / 1.30 (d, 6.5) / H-4 / C-3, C-4, C-5
18 / 17.2 / 0.98 (d, 6) / C-5, C-6, C-7
19 / 15.6 / 1.08 (d, 6.5) / H-8 / C-8, C-7, C-9
20 / 12.6 / 1.10 (d, 6.5) / H-12 / C-13, C-12, C-11
1 / 103.1 / 4.34 (d, 7) / H-2 / C-5
2 / 74.7 / 3.33 (m, 9, 6) / H-1, H-3 / C-1, C-3
3 / 76.9 / 3.46 (bt, 8.1) / H-2, H-4
4 / 75.5 / 3.16 (bt, 8.5) / H-3, H-5 / C-3
5 / 72.0 / 3.34 (m, 9, 6) / H-4, H-6 / C-3
6 / 17.8 / 1.30 (d, 6.5) / H-5 / C-5, C-4

Table S4NMR data of 3-O-demethyl-D-chalcosyl-narbonolide (ODDCNB) in CDCl3.

Position / C / H (m, J in Hz) / COSY / HMBC
1 / 169.6
2 / 50.5 / 3.83 (q, 7) / H-16 / C-1, C-3, C-16
3 / 209.1
4 / 48.3 / 2.93 (m) / H-5, H-17 / C-3, C-17
5 / 78.4 / 4.23 (brs) / H-4 / C-18
6 / 36.4 / 1.90 (overlapped) / H-7a, H-18
7a
7b / 37.6 / 1.47 (m)
1.19 (m) / H-6, H-8 / C-9
C-9
8 / 41.2 / 2.84 (m) / H-7a, H-19 / C-7, C-9, C-19
9 / 203.8
10 / 129.2 / 6.04 (d, 16) / H-11 / C-9, C-12
11 / 147.2 / 6.64 (dd, 16, 6) / H-10, H-12 / C-9, C-12, C-13, C-20
12 / 38.5 / 2.74 (m) / H-11, H-13, H-20 / C-10, C-11, C-13, C-20
13 / 79.1 / 4.94 (dt, 9.5, 3.5) / H-12, H-14a, H-14b
14a
14b / 22.8 / 1.59 (m)
1.50 (m) / H-13, H-15
H-13, H-15 / C-13
15 / 10.7 / 0.91 (t, 7.5) / H-14a, H-14b / C-13
16 / 14.3 / 1.37 (d, 7) / H-2 / C-1, C-2, C-3
17 / 13.8 / 1.32 (d, 7.5) / H-4 / C-3, C-4, C-5
18 / 17.1 / 1.01 (d, 7) / H-6 / C-5, C-6, C-7
19 / 15.4 / 1.10 (d, 7.5) / H-8 / C-8, C-7, C-9
20 / 12.9 / 1.11 (d, 7) / H-12 / C-13, C-12, C-11
1 / 103.3 / 4.31 (d, 7.5) / H-2 / C-5, C-3, C-5
2 / 76.9 / 3.25 (t, 8.5) / H-1, H-3 / C-1, C-3
3 / 71.4 / 3.70 (ddd, 14.5, 9, 5) / H-2, H-4a, H-4b / C-2
4a
4b / 39.9 / 1.95 (ddd, 13, 5, 1.5)
1.42 (m) / H-5, H-3
H-3 / C-2, C-3
C-3, C-5
5 / 68.4 / 3.62 (ddd, 11.5, 6.5, 1.5) / H-4, H-6 / C-6, C-1, C-3
6 / 21.1 / 1.25 (d, 6) / H-5 / C-5, C-4

1

Table S5 NMR data of L-rhamnosyl-narbonolide (LRHMNB) in CDCl3.

Position / C / H (m, J in Hz) / COSY / HMBC
1 / 169.6
2 / 50.3 / 3.81 (q, 7) / H-16 / C-1, C-3, C-16
3 / 208.4a
4 / 48.4 / 2.86 (m) / H-5, H-17
5 / 78.5a / 4.33 (brs) / H-4
6 / 36.5 / 1.80 (m) / H-7a, H-18
7a
7b / 37.3 / 1.46 (m)
1.19 (m) / H-6, H-8, H-7b
H-7a
8 / 41.7 / 2.83 (m) / H-7a, H-19
9 / 204.0a
10 / 129.1a / 6.04 (d, 16) / H-11 / C-9, C-12
11 / 147.4 / 6.62 (dd, 15.5, 5.5) / H-10, H-12
12 / 38.6 / 2.75 (m) / H-11, H-13, H-20
13 / 79.2 / 4.90 (brs) / H-12, H-14a, H-14b
14a
14b / 22.7 / 1.60 (m)
1.50 (m) / H-13, H-15
H-13, H-15
15 / 10.7 / 0.91 (t, 7) / H-14a, H-14b / C-13, C-14
16 / 14.3 / 1.35 (d, 6.5) / H-2 / C-1, C-2, C-3
17 / 13.5 / 1.25 (d, 7) / H-4 / C-3, C-4
18 / 17.1 / 1.00 (d,7) / H-6 / C-5, C-6, C-7
19 / 15.3 / 1.13 (d, 7) / H-8 / C-8, C-7, C-9
20 / 13.0 / 1.10 (d, 6.5) / H-12 / C-13, C-12, C-11
1 / 101.5 / 4.90 (brs) / H-2 / C-3, C-5, C-5
2 / 71.1 / 3.94 (brs) / H-1, H-3
3 / 71.9 / 3.73 (m, 9) / H-2, H-4
4 / 73.5 / 3.50 (m ) / H-5, H-3
5 / 68.8 / 3.73 (m, 9, 7) / H-4, H-6
6 / 17.6 / 1.30 (d, 6.5) / H-5 / C-5, C-4

a Assignments based on HSQC and HMBC spectra.

Table S6 NMR data of D-digitoxysyl-narbonolide (DDGTNB) in CDCl3.

Position / C / H (m, J in Hz) / COSY / HMBC
1 / 169.5
2 / 51.0 / 3.80 (q, 7.0) / H-16 / C-1, C-3, C-16
3 / 208.5a
4 / 49.0 / 2.84 (quin, 7.5) / H-5, H-17 / C-3, C-17, C-5, C-6
5 / 79.5 / 4.15 (brs) / H-4 / C-18
6 / 36.5 / 1.75 (m) / H-18
7a
7b / 37.0a / 1.44 (m)
1.19 (m) / H-8
H-8 / C-6, C-8, C-9, C-18, C-19
C-6, C-8, C-18, C-19
8 / 42.5 / 2.75 (m) / H-7a, H-7b, H-19 / C-7, C-9, C-19
9 / 203.0
10 / 129.0a / 6.12 (d, 16) / H-11 / C-9, C-12
11 / 147.0a / 6.68 (dd, 16, 6.5) / H-10, H-12 / C-9, C-12, C-20
12 / 38.6 / 2.73 (m) / H-11, H-20 / C-11, C-13
13 / 78.9 / 4.92 (dt, 9.5, 5.5) / H-14a
14a
14b / 23.0a / 1.60 (m)
1.54 (m) / H-15, H-13
H-15 / C-15
C-13, C-15
15 / 10.7 / 0.90 (t, 7.5) / H-14a, H-14b / C-13, C-14
16 / 14.6 / 1.37 (d, 7) / H-2 / C-1, C-2, C-3
17 / 14.0 / 1.23 (d, 7.5) / H-4 / C-3, C-4
18 / 17.2 / 1.01 (d, 7) / H-6 / C-5, C-6, C-7
19 / 15.8 / 1.13 (d, 7) / H-8 / C-8, C-7, C-9
20 / 13.0 / 1.10 (d, 7) / H-12 / C-13, C-12, C-11
1 / 98.9 / 4.84 (d, 9.5 ) / H-2b / C-5, C-2
2a
2b / 37.9 / 2.10 (dt, 13.5, 2)
1.70 (ddd, 13.7, 9.9, 2.2) / H-2b, H-3
H-1, 2a, H-3 / C-3, C-4
C-1
3 / 68.5 / 4.11 (m, 3) / H-2a, H-4, H-2b / C-1, C-5
4 / 73.2 / 3.32 (dd, 9.5, 3) / H-3, H-5 / C-6
5 / 69.5 / 3.72 (m, 9.5, 6.5) / H-4, H-6 / C-1, C-3, C-4, C-6
6 / 18.2 / 1.29 (d, 6.5) / H-5 / C-5, C-4

a Assignments based on HSQC and HMBC spectra.

Table S7 NMR data of D-boivinosyl-narbonolide (DBVNNB) in CDCl3.

Position / C / H (m, J in Hz) / COSY / HMBC
1 / 169.6
2 / 50.9 / 3.80 (q, 7.0) / H-16 / C-1, C-3, C-16
3 / 208.4
4 / 48.9 / 2.84 (quin, 7) / H-5, H-17 / C-3, C-5, C-6, C-17
5 / 79.0 / 4.17 (d like, 4) / H-4, H-6 / C-18
6 / 36.6 / 1.80 (m) / H-5, H-7b, H-18
7a
7b / 37.0 / 1.44 (m)
1.19 (m) / H-8
H-6 / C-9, C-8, C-6, C-18, C-19
C-9, C-5
8 / 42.2 / 2.78 (m, 6.5, 7) / H-19 / C-9, C-7, C-19
9 / 203.8
10 / 128.5a / 6.11 (d, 16) / H-11 / C-9, C-12
11 / 147.2 / 6.68 (dd, 15.5, 6) / H-10, H-12 / C-9, C-12, C-20
12 / 38.6 / 2.73 (m, 6.5, 2.5) / H-13, H-20 / C-11
13 / 79.0 / 4.92 (dt, 9.5, 4) / H-12, H-14a, H-14b
14a
14b / 22.9 / 1.59 (m)
1.52 (m) / H-13, H-15
H-13, H-15 / C-15, C-13
C-15, C-13
15 / 10.7 / 0.90 (t, 7) / H-14a, H-14b / C-13, C-14
16 / 14.5 / 1.36 (d, 7) / H-2 / C-1, C-2, C-3
17 / 13.9 / 1.22 (d, 7.5) / H-4 / C-3, C-4, C-5
18 / 17.2 / 1.00 (d, 7) / H-6 / C-5, C-6, C-7
19 / 15.7 / 1.12 (d, 7) / H-8 / C-8, C-7, C-9
20 / 12.8 / 1.10 (d, 7) / H-12 / C-13, C-12, C-11
1 / 99.7 / 4.81 (dd, 8, 4.5) / H-2a, H-2b / C-5, C-2
2 / 33.8 / 1.78 (dd, 8, 3)
1.80 (m) / H-1, H-3
H-1, H-3 / C-1, C-3
C-1, C-3
3 / 69.6 / 4.11 (m, 3) / H-2a, H-2b, H-4 / C-1
4 / 70.7 / 3.26 (brs) / H-3, H-5 / C-5, C-3
5 / 68.8 / 4.05 (m, 6.5) / H-4, H-6 / C-4, C-1, C-6
6 / 16.6 / 1.24 (d, 6.5) / H-5 / C-5, C-4

a Assignments based on HSQC and HMBC spectra.

Table S8 NMR data of L-olivosyl-narbonolide (LOLVNB) in CDCl3.

Position / C / H (m, J in Hz) / COSY / HMBC
1 / 169.6
2 / 50.4 / 3.82 (q, 7) / H-16 / C-1, C-3, C-16
3 / 208.5a
4 / 48.2a / 2.85 (m) / H-5, H-17 / C-3, C-6, C-17
5 / 77.5 a / 4.27 (brs) / H-4
6 / 36.5 / 1.80 (m) / H-18
7a
7b / 37.5a / 1.43 (m)
1.12 (m) / H-8 / C-6, C-9
8 / 42.0a / 2.82 (m) / H-19, H-7a / C-9
9 / 203.5
10 / 129.0 a / 6.06 (d, 16) / H-11 / C-9, C-12
11 / 147.0 / 6.63 (m) / H-10, H-12 / C-9
12 / 38.6 / 2.74 (m) / H-11, H-20
13 / 79.2 / 4.90 (m) / H-14a, H-14b
14a
14b / 22.5 / 1.60 (m)
1.50 (m) / H-13, H-15
H-13, H-15 / C-15
15 / 10.7 / 0.90 (t, 7) / H-14a, H-14b / C-13, C-14
16 / 14.4 / 1.37 (d, 7) / H-2 / C-1, C-2, C-3
17 / 13.9 / 1.24 (d, 7) / H-4 / C-3, C-4
18 / 17.2 / 1.00 (d, 7) / H-6 / C-5, C-6
19 / 15.2a / 1.13 (d, 6.5) / H-8 / C-8, C-7, C-9
20 / 13.0a / 1.10 (d, 7) / H-12 / C-12, C-11, C-13
1 / 98.9 / 4.97 (d, 3) / H-2a / C-3, C-5
2a
2b / 38.1 / 1.70 (td, 12.5, 4)
2.12 (dd, 13, 5) / H-3, H-2b, H-1
H-3, H-2a / C-3, C-4, C-1
C-3, C-4, C-1
3 / 69.4 / 3.90 (ddd, 12, 9, 5) / H-2a, H-2b, H-4 / C-4, C-5
4 / 78.5 / 3.13 (t, 9) / H-3, H-5 / C-3, C-5
5 / 6 8.5 / 3.73 (m, 9, 6) / H-4, H-6 / C-4
6 / 17.8 / 1.29 (d, 6) / H-5 / C-5, C-4

a Assignments based on HSQC and HMBC spectra.

Table S9 NMR data of L-digitoxosyl-narbonolide (LDGTNB) in CDCl3.

Position / C / H (m, J in Hz) / COSY / HMBC
1 / 169.5
2 / 50.1 / 3.80 (q, 7.0) / H-16 / C-1, C-3, C-16
3 / 208.1
4 / 48.5a / 2.85 (m) / H-5, H-17 / C-3, H-6, C-17
5 / 78.5 / 4.36 (brs) / H-4
6 / 36.6 / 1.82 (m) / H-7a, H-18
7a
7b / 37.2a / 1.48 (m)
1.10 (m) / H-6 / C-9, C-6, C-18, C-19
C-8, C-6, C-18, C-19
8 / 42.0a / 2.85 (m) / H-7b, H-7a, H-19 / C-9, C-7
9 / 203.2
10 / 129.0 / 6.02 (d, 16.5) / H-11 / C-9, C-12
11 / 147.4 / 6.63 (dd, 15.5, 5.5) / H-10, H-12 / C-9
12 / 38.6 / 2.74 (m) / H-11, H-13, H-20 / C-13, C-20
13 / 79.5 / 4.91 (dt, 10.5, 3.5) / H-12, H-14b, H-14a
14a
14b / 22.5a / 1.60 (m)
1.48 (m) / H-13, H-15, H-14b
H-14a, H-13, H-15 / C-13
15 / 10.7 / 0.90 (t, 7.5) / H-14a, H-14b / C-13, C-14
16 / 14.2 / 1.36 (d, 7.0) / H-2 / C-1, C-2, C-3
17 / 13.7 / 1.26 (d, 7.0) / H-4 / C-3, C-4
18 / 17.3 / 1.02 (d, 7.0) / H-6 / C-5, C-6, C-7
19 / 15.5 / 1.13 (d, 6.5) / H-8 / C-8, C-7, C-9
20 / 13.0a / 1.09 (d, 7.5) / H-12 / C-13, C-12, C-11
1 / 99.1 / 5.03 (brs, 3) / H-2a, 2b / C-3, C-5
2a
2b / 35.4 / 2.17 (dd, 14.5, 3.0)
1.91 (dt, 15.0, 3.5) / H-1, H-2b, H-3
H-1, H-2a, H-3 / C-1, C-3, C-4
3 / 67.5 / 3.96 (brs) / H-2a, H-2b, H-4 / C-5
4 / 72.9 / 3.15 (dd, 10, 3) / H-3, H-5 / C-5, C-6
5 / 65.5 / 3.83 (overlapped) / H-4, H-6 / C-3, C-4, C-6
6 / 17.9 / 1.33 (d, 6.5) / H-5 / C-4, C-5

a Assignments based on HSQC and HMBC spectra.

References

Hong JSJ, Park SH, Choi CY, Sohng JK, Yoon YJ(2004) New olivosyl derivatives of methymycin/pikromycin from an engineered strain of Streptomyces venezuelae. FEMS Microbiol Lett 238:291299

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