Indian Journal of Microbiology

, Volume 59, Issue 4, pp 436–444 | Cite as

Mutanase Enzyme from Paracoccus mutanolyticus RSP02: Characterization and Application as a Biocontrol Agent

  • Sudheer Kumar Buddana
  • Ravi Naga Amrutha
  • Uma Rajeswari Batchu
  • Suprasanna Penna
  • Reddy Shetty PrakashamEmail author
Original research article


Mutanases are enzymes that have the ability to cleave α-1,3 linkages in glucan polymer. In the present investigation, mutanase enzyme purified from the culture filtrate of Paracoccus mutanolyticus was evaluated for Streptococcal biofilm degradation and antimicrobial activity against pathogenic fungi along with enzyme kinetics, activation energies, pH and thermal stability. Biochemical and molecular characterization depicted that the enzyme showed optimum activity at pH 5.5 and at 50 °C. It displayed Michaelis–Menten behaviour with a Km of 1.263 ± 0.03 (mg/ml), Vmax of 2.712 ± 0.15 U/mg protein. Thermal stability studies denoted that it required 55.46 and 135.43 kJ mol−1 of energy for activation and deactivation in the temperature range of 30–50 °C and 50–70 °C respectively. Mutanase activity was enhanced ~ 50 and 75% by Fe2+ and EDTA, respectively, while presence of Hg2+ and Mn2+ inhibit > 90% of its activity. This enzyme has a molecular mass of 138 kDa and showed monomeric nature by Zymography. Scanning electron microscopy analysis of mutanase treated Streptococcal cells revealed cleavage of linkages among the cells and complete separation of cells, indicating its potential in dentistry as an anticaries agent in the prophylaxis and therapy of dental caries. In addition, antifungal activity of mutanase against Colletotrichum capsici MTCC 10147 and Cladosporium cladosporioide MTCC 7371 revealed that the enzyme has potential towards biological control of phytopathogens which could be used as an alternative bio-control agent against chemical pesticides in the future.


Mutanase Enzyme kinetics Streptococcal biofilm Zymography Antifungal activity 



Authors are thankful to the Director, CSIR-IICT Hyderabad. Mr. Sudheer Kumar B gratefully acknowledges the CSIR, New Delhi, for providing Senior Research Fellowship. R. Naga Amrutha thanks the Department of Science and Technology (DST), New Delhi for financial support. Uma Rajeswari Batchu for BRNS, Mumbai for providing JRF. The manuscript communication number through CSIR- IICT is IICT/Pubs/2019/223.

Supplementary material

12088_2019_821_MOESM1_ESM.pptx (715 kb)
Supplementary material 1 (PPTX 714 kb)


  1. 1.
    Wiater A, Pleszczynska M, Rogalski J, Szajnecka L, Szczodrak J (2013) Purification and properties of an α-(1 → 3)-glucanase (EC from Trichoderma harzianum and its use for reduction of artificial dental plaque accumulation. Acta Biochim Pol 60:123–128CrossRefGoogle Scholar
  2. 2.
    Pleszczynska M, Wiater A, Szczodrak J (2010) Mutanase from Paenibacillus sp. MP-1 produced inductively by fungal α-1,3-glucan and its potential for the degradation of mutan and Streptococcus mutans biofilm. Biotechnol Lett 32:1699–1704. CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Sanz L, Montero M, Redondo J, Llobell A, Monte E (2005) Expression of an α-1,3-glucanase during mycoparasitic interaction of Trichoderma asperellum. FEBS J 272:493–499. CrossRefPubMedGoogle Scholar
  4. 4.
    Wiater A, Pleszynska M, Pietrykowska-Tudruj E, Janczarek M, Staniec B, Szczodrak J (2019) Isolation and characterization of α-(1-3)-glucan-degrading bacteria from the gut of Diaperis boleti feeding on Laetiporus sulphurous. Entomol Sci 22:36–41. CrossRefGoogle Scholar
  5. 5.
    Zonneveld BJM (1972) A new type of enzyme, an exo-splitting α-1,3 glucanase from non-induced cultures of Aspergillus nidulans. Biochem Biophys Acta 258:541–547. CrossRefPubMedGoogle Scholar
  6. 6.
    Buddana SK, Shetty PR, Krothapalli SRRS (2016) An endolytic mutanase from novel strain Paracoccus mutanolyticus: it’s application potential in dentistry. J Med Microbiol 65:985–991. CrossRefPubMedGoogle Scholar
  7. 7.
    Matsuda S, Kawanami Y, Takeda H, Ooi T, Kinoshita S (1997) Purification and properties of mutanase from Bacillus circulans. J Ferment Bioeng 83:593–595. CrossRefGoogle Scholar
  8. 8.
    Takehara T, Inoue M, Morioka T, Yokogawa K (1981) Purification and properties of endo-alpha-1,3-glucanase from a Streptomyces charteuses strain. J Bacteriol 145:729–735PubMedPubMedCentralGoogle Scholar
  9. 9.
    Suyotha W, Fujiki H, Cherdvorapong V, Takagi K, Yano S, Wakayama M (2017) A novel thermostable a-1,3-glucanase from Streptomyces thermodiastaticus HF 3-3. J Gen Appl Microbiol 63:296–304. CrossRefPubMedGoogle Scholar
  10. 10.
    Cherdvorapong V, Fujiki H, Suyotha W, Yoichi T, Shigekazu Y, Kazuyoshi T, Mamoru W (2019) Enzymatic and molecular characterization of α-1,3-glucanase (AglST2) from Streptomyces thermodiastaticus HF3-3 and its relation with α-1,3-glucanase HF65 (AglST1). J Gen Appl Microbiol 65:18–25. CrossRefPubMedGoogle Scholar
  11. 11.
    Wiater A, Szczodrak J, Pleszczynska M, Prochniak K (2005) Production and use of mutanase from Trichoderma harzianum for effective degradation of Streptococcal mutans. Braz J Microbiol 36:137–146. CrossRefGoogle Scholar
  12. 12.
    Inoue M, Yakushiji T, Katsuki M, Kudo N, Koga T (1988) Reduction of the adherence of Streptococcus sobrinus insoluble α-d-glucan by endo-(1→3)-α-d-glucanase. Carbohydr Res 182:277–286. CrossRefPubMedGoogle Scholar
  13. 13.
    Buddana SK, Yashwanth VV, Prakasham RS (2015) Fibrinolytic, anti-inflammatory and anti-microbial properties of a-(1-3)-glucans produced from Streptococcus mutans (MTCC 497). Carbohydr Polym 115:152–159. CrossRefPubMedGoogle Scholar
  14. 14.
    Somogyi M (1952) Notes on sugar determination. J Biol Chem 195:19–23Google Scholar
  15. 15.
    Nelson N (1944) A photometric adaptation of the Somogyi method for the determination of glucose. J Biol Chem 153:375–380Google Scholar
  16. 16.
    Wiater A, Szczodrak J, Rogalski J (2001) Purification and characterization of an extracellular mutanase from Trichoderma harzianum. Mycol Res 105:1357–1363. CrossRefGoogle Scholar
  17. 17.
    Otsuka R, Imai S, Murata T, Nomur Y, Okamoto M, Tsumori H, Kakuta E, Hanada N, Momoi Y (2015) Application of chimeric glucanase comprising mutanase and dextranase for prevention of dental biofilm formation. Microbiol Immunol 59:28–36. CrossRefPubMedGoogle Scholar
  18. 18.
    Pleszczynska M, Wiater A, Skowronek M, Szczodrak J (2012) Purification and characterization of mutanase produced by Paenibacillus curdlanolyticus MP-1. Prep Biochem Biotechnol 42:335–347CrossRefGoogle Scholar
  19. 19.
    Pleszczynska M, Wiater A, Janczarek M, Szczodrak J (2015) (1→3)-α-d-Glucan hydrolases in dental biofilm prevention and control: a review. Int J Biol Macromol 79:761–778. CrossRefPubMedGoogle Scholar
  20. 20.
    Suyotha W, Yano S, Itoh T, Fujimoto H, Hibi T, Tachiki T, Wakayama M (2014) Characterization of α-1,3-glucanase isozyme from Paenibacillus glycanilyticus FH11 in a new subgroup of family 87 α-1,3-glucanase. J Biosci Bioeng 118:378–385. CrossRefPubMedGoogle Scholar
  21. 21.
    Meyer MT, Phaff HG (1980) Purification and properties of (1→3)-α-glucanases from Bacillus circulans WL-12. J Gen Microbiol 118:197–208. CrossRefGoogle Scholar
  22. 22.
    Sumitomo N, Saeki K, Ozaki K, Ito S, Kobayashi T (2007) Mutanase from a Paenibacillus isolate: nucleotide sequence of the gene and properties of recombinant enzymes. Biochem Biophys Acta 1770:716–724. CrossRefPubMedGoogle Scholar
  23. 23.
    Riordan JF (1977) The role of metals in enzyme activity. Ann Clin Lab Sci 7:119–129PubMedGoogle Scholar
  24. 24.
    Simonson LG, Gaugler RW, Lamberts BL, Reiher DA (1982) Purification and properties of endo-1,3-a-D-glucanase from Pseudomonas. Biochem Biophys Acta 715:189–195. CrossRefPubMedGoogle Scholar
  25. 25.
    Wiater A, Szczodrak J, Rogalski J (2004) Hydrolysis of mutan and prevention of its formation in streptococcal films by fungal α-d-glucanases. Process Biochem 39:1481–1489. CrossRefGoogle Scholar
  26. 26.
    Fujikawa T, Sakaguchi A, Nishizawa Y, Kouzai Y, Minami E (2012) Surface α-1, 3-glucan facilitates fungal stealth infection by interfering with innate immunity in plants. PLoS Pathog 8:1–16. CrossRefGoogle Scholar
  27. 27.
    Zarei M, Saeed A, Hossein Z, Alireza S, Morteza D, Kambiz NA, Ahmad G, Abbasali M (2011) Characterization of a chitinase with antifungal activity from a native Serratia marcescens B4A. Braz J Microbiol 42:1017–1029. CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Bachtiar EW, Bachtiar BM (2018) Relationship between Candida albicans and Streptococcus mutans in early childhood caries, evaluated by quantitative PCR [version 2; Peer review: 2 approved]. F1000Research 7:1645CrossRefGoogle Scholar
  29. 29.
    Ait-Lahsen H, Soler A, Rey M, de la Cruz J, Monte E, Llobell A (2001) An antifungal exo-α-1,3-glucanase (AGN13.1) from the biocontrol fungus Trichoderma harzianum. App Environ Microbiol 67:5833–5839. CrossRefGoogle Scholar

Copyright information

© Association of Microbiologists of India 2019

Authors and Affiliations

  1. 1.Medicinal Chemistry and BiotechnologyCSIR-Indian Institute of Chemical TechnologyHyderabadIndia
  2. 2.Academy of Scientific and Innovative Research (AcSIR)CSIR-Indian Institute of Chemical TechnologyHyderabadIndia
  3. 3.Nuclear Agriculture and Biotechnology DivisionBhabha Atomic Research Centre (BARC)MumbaiIndia

Personalised recommendations