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Inhibitory Effects of Streptomyces sp. MBTH32 Metabolites on Sortase A and Sortase A-Mediated Cell Clumping of Staphylococcus aureus to Fibrinogen
1Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National University, Seoul 08826, Republic of Korea, 2Natural Products Research Institute, College of Pharmacy, Seoul National University, Seoul 08826, Republic of Korea
Correspondence to:J. Microbiol. Biotechnol. 2019; 29(10): 1603-1606
Published October 28, 2019 https://doi.org/10.4014/jmb.1906.06026
Copyright © The Korean Society for Microbiology and Biotechnology.
Abstract
Keywords
Body
The ocean environment covers 70% of the Earth’s surface and has more diverse conditions than the terrestrial environment, such as low oxygen and lack of light [1, 2]. Under these conditions, biosynthesis of secondary metabolites typically involves mechanisms modified for physiological adaptation, which increases the probability that unusual natural products might be present [3, 4]. In the last decade, numerous natural products have been discovered in the marine environment; notably, thousands of those compounds exhibit new structures, and three-fourths of them demonstrate diverse bioactivities [5]. Marine-derived
Sortases, transpeptidases that anchor surface proteins to the peptidoglycan layer in gram-positive bacteria, have attracted attention as a potential target of novel antibiotics [10]. Surface proteins covalently tethered to the cell wall by sortases allow gram-positive bacteria to adhere to host tissues and to invade epithelial cells [11, 12]. Sortases comprise six isoforms, A–F (SrtA–F). Among them, SrtA has been shown to play an important role in the pathogenesis of
In our search for SrtA inhibitors in marine-derived
Strain MBTH32 was isolated from marine sediment from Shinjin Island, Republic of Korea; it showed 98% identity to
-
Fig. 1. Structures of compounds
1–3 isolated from marinederivedStreptomyces sp. MBTH32: enterocin (1 ),N -acetyl-β-oxotryptamine (2 ), and lumichrome (3 ).
Recombinant SrtA was purified from transformed
-
Table 1 . Inhibitory effects of compounds
1–3 on the activity of SrtA and bacterial growth ofS. aureus ATCC6538p.Compound IC50 (μM) MIC (μM) SrtA S. aureus ATCC6538p1 594.76 ± 3.78 9.00 2 1108.65 ± 7.52 >1185.19 3 198.20 ± 0.94 >528.42 Berberine chloride 106.40 ± 1.36 >344.26 Berberine chloride was used as a reference inhibitor of SrtA. IC50 values are the mean ± SD (
n = 3).
-
Fig. 2. Lineweaver-Burk plot of SrtA inhibition by compound
3 . Compound3 was applied at IC50 and at 2× IC50 concentrations. [S], reaction substrate concentration; V, reaction velocity (Δfluorescence/ min). Each point indicates the mean ± SD of three independent experiments.
SrtA has been reported to immobilize fibrinogen-binding protein, thus accelerating bacterial adhesion to host tissues and subsequent invasion [28, 29]. We hypothesized that the immobilization of fibrinogen-binding protein may be reduced by suppression of SrtA activity
-
Fig. 3. Effects of
srtA gene expression and SrtA inhibitors on the clumping ofS. aureus with fibrinogen. (A ) Clumping assay was performed withS. aureus Newman (wild-type) and SKM12 (srtA -knockout mutant) strains. (B ) Berberine chloride and compound3 were applied at the indicated concentrations at 37°C for 2 h. Thet -test was used for statistical analysis of multiple comparisons. A value ofp < 0.05 was used as the criterion for statistical significance. *p < 0.05, **p < 0.01, ***p < 0.001.
In conclusion, three metabolites isolated from marine-derived
Acknowledgments
This research was supported by the Basic Science Research Program through the National Research Foundation (NRF-2018R1D1A1B07043375) of Korea funded by the Ministry of Education, Science and Technology.
Conflict of Interest
The authors have no financial conflicts of interest to declare.
References
- Tortorella E, Tedesco P, Palma Esposito F, January G, Fani R, Jaspars M,
et al . 2018. Antibiotics from deep-sea microorganisms:current discoveries and perspectives.Mar. Drugs 16 . pii:E355. - Tyler PA. 2003.
Ecosystems of the deep oceans , pp. 1-4. 1st Ed. Elsevier Science, Amsterdam. - Wright PC, Westacott RE, Burja AM. 2003. Piezotolerance as a metabolic engineering tool for the biosynthesis of natural products.
Biomol. Eng. 20 : 325-331. - Bull AT, Ward AC, Goodfellow M. 2000. Search and discovery strategies for biotechnology: the paradigm shift.
Microbiol. Mol. Biol. Rev. 64 : 573-606. - Blunt JW, Carroll AR, Copp BR, Davis RA, Keyzers RA, Prinsep MR. 2018. Marine natural products.
Nat. Prod. Rep. 35 : 8-53. - Braña AF, Sarmiento-Vizcaíno A, Pérez-Victoria I, Otero L, Fernández J, Palacios JJ,
et al . 2017. Branimycins B and C, antibiotics produced by the abyssal actinobacteriumPseudonocardia carboxydivorans M-227.J. Nat. Prod. 80 : 569-573. - Le TC, Yang I, Yoon YJ, Nam SJ, Fenical W. 2016. Ansalactams B-D illustrate further biosynthetic plasticity within the ansamycin pathway.
Org. Lett. 18 : 2256-2259. - Song Y, Li Q, Liu X, Chen Y, Zhang Y, Sun A,
et al . 2014. Cyclic hexapeptides from the deep South China Sea-derivedStreptomyces scopuliridis SCSIO ZJ46 active against pathogenic gram-positive bacteria.J. Nat. Prod. 77 : 1937-1941. - Pan HQ, Zhang SY, Wang N, Li ZL, Hua HM, Hu JC,
et al . 2013. New spirotetronate antibiotics, lobophorins H and I, from a South China Sea-derivedStreptomyces sp. 12A35.Mar. Drugs 11 : 3891-3901. - Cossart P, Jonquières R. 2000. Sortase, a universal target for therapeutic agents against Gram-positive bacteria? Proc.
Natl. Acad. Sci. USA 97 : 5013-5015. - Mazmanian SK, Liu G, Jensen ER, Lenoy E, Schneewind O. 2000.
Staphylococcus aureus sortase mutants defective in the display of surface proteins and in the pathogenesis of animal infections.Proc. Natl. Acad. Sci. USA 97 : 5510-5515. - Wesson CA, Liou LE, Todd KM, Bohach GA, Trumble WR, Bayles KW. 1998.
Staphylococcus aureus A grand Sar global regulators influence internalization and induction of apoptosis.Infect. Immun. 66 : 5238-5243. - Mazmanian SK, Ton-That H, Su K, Schneewind O. 2002. An iron-regulated sortase anchors a class of surface protein during
Staphylococcus aureus pathogenesis.Proc. Natl. Acad. Sci. USA 99 : 2293-2298. - Mazmanian SK, Skaar EP, Gaspar AH, Humayun M, Gornicki P, Jelenska J,
et al . 2003. Passage of heme-iron across the envelope ofStaphylococcus aureus .Science 299 : 906-909. - Frankel BA, Bentley M, Kruger RG, McCafferty DG. 2004. Vinyl sulfones: inhibitors of SrtA, a transpeptidase required for cell wall protein anchoring and virulence in
Staphylococcus aureus .J. Am. Chem. Soc. 126 : 3404-3405. - Sibbald MJ, Yang XM, Tsompanidou E, Qu D, Hecker M, Becher D,
et al . 2012. Partially overlapping substrate specificities of staphylococcal group A sortases.Proteomics 12 : 3049-3062. - Cascioferro S, Raffa D, Maggio B, Raimondi MV, Schillaci D, Daidone G. 2015. Sortase A inhibitors: recent advances and future perspectives.
J. Med. Chem. 58 : 9108-9123. - Kang H, Jensen PR, Fenical W. 1996. Isolation of microbial antibiotics from a marine ascidian of the genus Didemnum.
J. Org. Chem. 61 : 1543-1546. - Martínez-Luis S, Gómez JF, Spadafora C, Guzmán HM, Gutiérrez M. 2012. Antitrypanosomal alkaloids from the marine bacterium
Bacillus pumilus .Molecules 17 : 11146-11155. - Andrioli WJ, Lopes AA, Cavalcanti BC, Pessoa C, Nanayakkara NPD, Bastos JK. 2017. Isolation and characterization of 2pyridone alkaloids and alloxazines from
Beauveria bassiana .Nat. Prod. Res. 31 : 1920-1929. - Lee KY, Shin DS, Yoon JM, Kang HJ, Oh KB. 2002. Expression of sortase, a transpeptidase for cell wall sorting reaction, from
Staphylococcus aureus ATCC 6538p inEscherichia coli .J. Microbiol. Biotechnol. 12 : 530-533. - Oh KB, Kim SH, Lee J, Cho WJ, Lee T, Kim S. 2004. Discovery of diarylacrylonitriles as a novel series of small molecule sortase A inhibitors.
J. Med. Chem. 47 : 2418-2421. - Hu P, Huang P, Chen WM. 2013. Curcumin inhibits the sortase A activity of the
Streptococcus mutans UA159.Appl. Biochem. Biotechnol. 171 : 396-402. - Kim SH, Shin DS, Oh MN, Chung SC, Lee JS, Oh KB. 2004. Inhibition of the bacterial surface protein anchoring transpeptidase sortase by isoquinoline alkaloids.
Biosci. Biotechnol. Biochem. 68 : 421-424. - Jonsson M, Mazmanian SK, Schneewind O, Bremell T, Tarkowski A. 2003. The role of
Staphylococcus aureus sortase A and sortase B in murine arthritis.Microbes Infect. 5 : 775-780. - Weinstein MP, Limbago B, Patel JB, Mathers AJ, Burnham C, Mazzulli T,
et al . 2018.Methods for Dilution Antimicrobial Susceptibility Test for Bacteria That Grow aerobically , pp. 15-50. 11th Ed. Clinical and Laboratory Standards Institute, Wayne, Pennsylvania. - Lineweaver H, Burk D. 1934. The determination of enzyme dissociation constants.
J. Am. Chem. Soc. 56 : 658-666. - Alksne LE, Projan SJ. 2000. Bacterial virulence as a target for antimicrobial chemotherapy.
Curr. Opin. Biotechnol. 11 : 625-636. - Zhang B, Teng Z, Li X, Lu G, Deng X, Niu X, Wang J. 2017. Chalcone attenuates
Staphylococcus aureus virulence by targeting sortase A and alpha-hemolysin.Front. Microbiol. 8 : 1715. - Weiss WJ, Lenoy E, Murphy T, Tardio L, Burgio P, Projan SJ,
et al . 2004. Effect of srtA and srtB gene expression on the virulence ofStaphylococcus aureus in animal models of infection.J. Antimicrob. Chemother. 53 : 480-486.
Related articles in JMB

Article
Note
J. Microbiol. Biotechnol. 2019; 29(10): 1603-1606
Published online October 28, 2019 https://doi.org/10.4014/jmb.1906.06026
Copyright © The Korean Society for Microbiology and Biotechnology.
Inhibitory Effects of Streptomyces sp. MBTH32 Metabolites on Sortase A and Sortase A-Mediated Cell Clumping of Staphylococcus aureus to Fibrinogen
Beomkoo Chung 1, Oh-Seok Kwon 2, Jongheon Shin 2* and Ki-Bong Oh 1*
1Department of Agricultural Biotechnology, College of Agriculture and Life Sciences, Seoul National University, Seoul 08826, Republic of Korea, 2Natural Products Research Institute, College of Pharmacy, Seoul National University, Seoul 08826, Republic of Korea
Correspondence to:Jongheon Shin shinj@snu.ac.kr
Ki-Bong Oh ohkibong@snu.ac.kr
Abstract
Sortase A (SrtA), a type of transpeptidase responsible for anchoring surface proteins to the peptidoglycan cell wall, is important in the virulence of gram-positive bacteria. Three compounds were isolated from marine-derived Streptomyces sp. MBTH32 using various chromatography techniques. The structures of these compounds were determined based on spectroscopic data and comparisons with previously reported data. Among the metabolites tested, lumichrome showed strong inhibitory activity against Staphylococcus aureus SrtA without affecting cell viability. The results of cell clumping activity assessment suggest the potential for using this compound to treat S. aureus infection by inhibiting SrtA activity.
Keywords: Marine Streptomyces, lumichrome, Staphylococcus aureus, sortase A, cell clumping
Body
The ocean environment covers 70% of the Earth’s surface and has more diverse conditions than the terrestrial environment, such as low oxygen and lack of light [1, 2]. Under these conditions, biosynthesis of secondary metabolites typically involves mechanisms modified for physiological adaptation, which increases the probability that unusual natural products might be present [3, 4]. In the last decade, numerous natural products have been discovered in the marine environment; notably, thousands of those compounds exhibit new structures, and three-fourths of them demonstrate diverse bioactivities [5]. Marine-derived
Sortases, transpeptidases that anchor surface proteins to the peptidoglycan layer in gram-positive bacteria, have attracted attention as a potential target of novel antibiotics [10]. Surface proteins covalently tethered to the cell wall by sortases allow gram-positive bacteria to adhere to host tissues and to invade epithelial cells [11, 12]. Sortases comprise six isoforms, A–F (SrtA–F). Among them, SrtA has been shown to play an important role in the pathogenesis of
In our search for SrtA inhibitors in marine-derived
Strain MBTH32 was isolated from marine sediment from Shinjin Island, Republic of Korea; it showed 98% identity to
-
Figure 1. Structures of compounds
1–3 isolated from marinederivedStreptomyces sp. MBTH32: enterocin (1 ),N -acetyl-β-oxotryptamine (2 ), and lumichrome (3 ).
Recombinant SrtA was purified from transformed
-
Table 1 . Inhibitory effects of compounds
1–3 on the activity of SrtA and bacterial growth ofS. aureus ATCC6538p..Compound IC50 (μM) MIC (μM) SrtA S. aureus ATCC6538p1 594.76 ± 3.78 9.00 2 1108.65 ± 7.52 >1185.19 3 198.20 ± 0.94 >528.42 Berberine chloride 106.40 ± 1.36 >344.26 Berberine chloride was used as a reference inhibitor of SrtA. IC50 values are the mean ± SD (
n = 3)..
-
Figure 2. Lineweaver-Burk plot of SrtA inhibition by compound
3 . Compound3 was applied at IC50 and at 2× IC50 concentrations. [S], reaction substrate concentration; V, reaction velocity (Δfluorescence/ min). Each point indicates the mean ± SD of three independent experiments.
SrtA has been reported to immobilize fibrinogen-binding protein, thus accelerating bacterial adhesion to host tissues and subsequent invasion [28, 29]. We hypothesized that the immobilization of fibrinogen-binding protein may be reduced by suppression of SrtA activity
-
Figure 3. Effects of
srtA gene expression and SrtA inhibitors on the clumping ofS. aureus with fibrinogen. (A ) Clumping assay was performed withS. aureus Newman (wild-type) and SKM12 (srtA -knockout mutant) strains. (B ) Berberine chloride and compound3 were applied at the indicated concentrations at 37°C for 2 h. Thet -test was used for statistical analysis of multiple comparisons. A value ofp < 0.05 was used as the criterion for statistical significance. *p < 0.05, **p < 0.01, ***p < 0.001.
In conclusion, three metabolites isolated from marine-derived
Acknowledgments
This research was supported by the Basic Science Research Program through the National Research Foundation (NRF-2018R1D1A1B07043375) of Korea funded by the Ministry of Education, Science and Technology.
Conflict of Interest
The authors have no financial conflicts of interest to declare.
Fig 1.

Fig 2.

Fig 3.

-
Table 1 . Inhibitory effects of compounds
1–3 on the activity of SrtA and bacterial growth ofS. aureus ATCC6538p..Compound IC50 (μM) MIC (μM) SrtA S. aureus ATCC6538p1 594.76 ± 3.78 9.00 2 1108.65 ± 7.52 >1185.19 3 198.20 ± 0.94 >528.42 Berberine chloride 106.40 ± 1.36 >344.26 Berberine chloride was used as a reference inhibitor of SrtA. IC50 values are the mean ± SD (
n = 3)..
References
- Tortorella E, Tedesco P, Palma Esposito F, January G, Fani R, Jaspars M,
et al . 2018. Antibiotics from deep-sea microorganisms:current discoveries and perspectives.Mar. Drugs 16 . pii:E355. - Tyler PA. 2003.
Ecosystems of the deep oceans , pp. 1-4. 1st Ed. Elsevier Science, Amsterdam. - Wright PC, Westacott RE, Burja AM. 2003. Piezotolerance as a metabolic engineering tool for the biosynthesis of natural products.
Biomol. Eng. 20 : 325-331. - Bull AT, Ward AC, Goodfellow M. 2000. Search and discovery strategies for biotechnology: the paradigm shift.
Microbiol. Mol. Biol. Rev. 64 : 573-606. - Blunt JW, Carroll AR, Copp BR, Davis RA, Keyzers RA, Prinsep MR. 2018. Marine natural products.
Nat. Prod. Rep. 35 : 8-53. - Braña AF, Sarmiento-Vizcaíno A, Pérez-Victoria I, Otero L, Fernández J, Palacios JJ,
et al . 2017. Branimycins B and C, antibiotics produced by the abyssal actinobacteriumPseudonocardia carboxydivorans M-227.J. Nat. Prod. 80 : 569-573. - Le TC, Yang I, Yoon YJ, Nam SJ, Fenical W. 2016. Ansalactams B-D illustrate further biosynthetic plasticity within the ansamycin pathway.
Org. Lett. 18 : 2256-2259. - Song Y, Li Q, Liu X, Chen Y, Zhang Y, Sun A,
et al . 2014. Cyclic hexapeptides from the deep South China Sea-derivedStreptomyces scopuliridis SCSIO ZJ46 active against pathogenic gram-positive bacteria.J. Nat. Prod. 77 : 1937-1941. - Pan HQ, Zhang SY, Wang N, Li ZL, Hua HM, Hu JC,
et al . 2013. New spirotetronate antibiotics, lobophorins H and I, from a South China Sea-derivedStreptomyces sp. 12A35.Mar. Drugs 11 : 3891-3901. - Cossart P, Jonquières R. 2000. Sortase, a universal target for therapeutic agents against Gram-positive bacteria? Proc.
Natl. Acad. Sci. USA 97 : 5013-5015. - Mazmanian SK, Liu G, Jensen ER, Lenoy E, Schneewind O. 2000.
Staphylococcus aureus sortase mutants defective in the display of surface proteins and in the pathogenesis of animal infections.Proc. Natl. Acad. Sci. USA 97 : 5510-5515. - Wesson CA, Liou LE, Todd KM, Bohach GA, Trumble WR, Bayles KW. 1998.
Staphylococcus aureus A grand Sar global regulators influence internalization and induction of apoptosis.Infect. Immun. 66 : 5238-5243. - Mazmanian SK, Ton-That H, Su K, Schneewind O. 2002. An iron-regulated sortase anchors a class of surface protein during
Staphylococcus aureus pathogenesis.Proc. Natl. Acad. Sci. USA 99 : 2293-2298. - Mazmanian SK, Skaar EP, Gaspar AH, Humayun M, Gornicki P, Jelenska J,
et al . 2003. Passage of heme-iron across the envelope ofStaphylococcus aureus .Science 299 : 906-909. - Frankel BA, Bentley M, Kruger RG, McCafferty DG. 2004. Vinyl sulfones: inhibitors of SrtA, a transpeptidase required for cell wall protein anchoring and virulence in
Staphylococcus aureus .J. Am. Chem. Soc. 126 : 3404-3405. - Sibbald MJ, Yang XM, Tsompanidou E, Qu D, Hecker M, Becher D,
et al . 2012. Partially overlapping substrate specificities of staphylococcal group A sortases.Proteomics 12 : 3049-3062. - Cascioferro S, Raffa D, Maggio B, Raimondi MV, Schillaci D, Daidone G. 2015. Sortase A inhibitors: recent advances and future perspectives.
J. Med. Chem. 58 : 9108-9123. - Kang H, Jensen PR, Fenical W. 1996. Isolation of microbial antibiotics from a marine ascidian of the genus Didemnum.
J. Org. Chem. 61 : 1543-1546. - Martínez-Luis S, Gómez JF, Spadafora C, Guzmán HM, Gutiérrez M. 2012. Antitrypanosomal alkaloids from the marine bacterium
Bacillus pumilus .Molecules 17 : 11146-11155. - Andrioli WJ, Lopes AA, Cavalcanti BC, Pessoa C, Nanayakkara NPD, Bastos JK. 2017. Isolation and characterization of 2pyridone alkaloids and alloxazines from
Beauveria bassiana .Nat. Prod. Res. 31 : 1920-1929. - Lee KY, Shin DS, Yoon JM, Kang HJ, Oh KB. 2002. Expression of sortase, a transpeptidase for cell wall sorting reaction, from
Staphylococcus aureus ATCC 6538p inEscherichia coli .J. Microbiol. Biotechnol. 12 : 530-533. - Oh KB, Kim SH, Lee J, Cho WJ, Lee T, Kim S. 2004. Discovery of diarylacrylonitriles as a novel series of small molecule sortase A inhibitors.
J. Med. Chem. 47 : 2418-2421. - Hu P, Huang P, Chen WM. 2013. Curcumin inhibits the sortase A activity of the
Streptococcus mutans UA159.Appl. Biochem. Biotechnol. 171 : 396-402. - Kim SH, Shin DS, Oh MN, Chung SC, Lee JS, Oh KB. 2004. Inhibition of the bacterial surface protein anchoring transpeptidase sortase by isoquinoline alkaloids.
Biosci. Biotechnol. Biochem. 68 : 421-424. - Jonsson M, Mazmanian SK, Schneewind O, Bremell T, Tarkowski A. 2003. The role of
Staphylococcus aureus sortase A and sortase B in murine arthritis.Microbes Infect. 5 : 775-780. - Weinstein MP, Limbago B, Patel JB, Mathers AJ, Burnham C, Mazzulli T,
et al . 2018.Methods for Dilution Antimicrobial Susceptibility Test for Bacteria That Grow aerobically , pp. 15-50. 11th Ed. Clinical and Laboratory Standards Institute, Wayne, Pennsylvania. - Lineweaver H, Burk D. 1934. The determination of enzyme dissociation constants.
J. Am. Chem. Soc. 56 : 658-666. - Alksne LE, Projan SJ. 2000. Bacterial virulence as a target for antimicrobial chemotherapy.
Curr. Opin. Biotechnol. 11 : 625-636. - Zhang B, Teng Z, Li X, Lu G, Deng X, Niu X, Wang J. 2017. Chalcone attenuates
Staphylococcus aureus virulence by targeting sortase A and alpha-hemolysin.Front. Microbiol. 8 : 1715. - Weiss WJ, Lenoy E, Murphy T, Tardio L, Burgio P, Projan SJ,
et al . 2004. Effect of srtA and srtB gene expression on the virulence ofStaphylococcus aureus in animal models of infection.J. Antimicrob. Chemother. 53 : 480-486.