Impact of cultivation conditions on xylanase production and growth in Paenibacillus mucilaginosus

Xylanase is an enzyme that hydrolyses β-1,4 bonds in plant xylan. This enzyme is applied in the bioconversion of agro-industrial waste for xylooligosaccharide hydrolysate production to improve digestibility and nutrition value of animal feed, food processing, the utilisation and faster decomposition of crop debris in soil, as well as in cellulose bleaching and other industries. The current trend focuses on using renewable resources, such as agricultural waste, as substitutes for expensive purified xylan in producer screening and xylanase synthesis. This work aimed to determine the impact of Paenibacillus mucilaginosus cultivation conditions on the xylanase production yield. Rice bran ferment lysate along with birch and beech timber xylans were used as a carbon source. Temperature, medium pH, pH correction factors, inoculant incubation time, carbon and nitrogen sources and concentrations were the studied criteria of xylanase biosynthesis and growth in bacteria P. ucilaginosus strain 560. We show that the xylanase biosynthesis and cultivation in P. mucilaginosus strain 560 are more practical and cost-effective with the use of a rice bran ferment lysate-based nutrient medium. Inductors contained in the rice bran ferment lysate improve the xylanase biosynthesis. Calcium ions also facilitate biosynthesis in the studied strain. Cultivation recommendations are: carbon source concentration in medium 0.5% of total reducing substances content; 0.2% carbamide as optimal nitrogen source; calcium hydroxide as an agent for medium pH correction to 6.0±0.2; cultivation temperature 30±1 °С. Under the specified conditions, cultivation of P. mucilaginosus does not require inoculate preprocessing, and a maximal xylanase activity in stationary culture reaches 20 U/mL.

1. Collins T, Gerday C, Feller G. Xylanases, xylanase families and extremophilic xylanases. FEMS Microbiology Reviews. 2005;29(1):3–23. https://doi.org/10.1016/j.femsre.2004.06.005

2. Sedlmeyer FB. Xylan as a by-product of biorefineries: characteristics and potential use for food applications. Food Hydrocolloids. 2014;25(8):1891–1898. https://doi.org/10.1016/J.FOODHYD.2011.04.005

3. Shanthi V, Roymon MG. Isolation and screening of alkaline thermostable xylanase producing bacteria from soil in Bhilai Durg region of Chhattisgarh, India. International Journal of Current Microbiology and Applied Sciences. 2014;3(8):303–311.

4. Habibi Y, Vignon MR. Isolation and characterrization of xylans from seed pericarp of Argania spinosa fruit. Carbohydrate Research. 2005;340(7):14 31–1436. https://doi.org/10.1016/j.carres.2005.01.039

5. Singh S, Madlala AM, Prior BA. Thermomyces lanuginosus: properties of strains and their hemicellulases. FEMS Microbiology Reviews. 2003;27 (1):3–16. https://doi.org/10.1016/S0168-6445(03)00018-4

6. Kulkarni N, Shendye A, Rao M. Molecular and biotechnological aspects of xylanases. FEMS Microbiology reviews. 1999;23(4):411–456. https://doi.org/10.1111/j.1574-6976.1999.tb00407.x

7. Fengel D, Wegener G. Wood: Chemistry. Ultrastructure. Reactions. Berlin. Walter de Gruyter, 1984 (Russ ed.: Fengel D, Vegener G. Drevesina. Khimiya. Ul’trastruktura. Reaktsii. Moscow: Lesnaya promyshlennost': 1988. 511 p.)

8. Chanda SK, Hirst EL, Jones JKN, Percival EGV. 262. The constitution of xylan from esparto grass (Stipa tenacissima, L.). Journal of the Chemical Society (Resumed). 1950:1289–1297. https://doi.org/10.1039/JR9500001289

9. Eda S, Ohnishi A, Katō K. Xylan isolated from the stalk of Nicotiana tabacum. Agricultural and Biological Chemistry. 1976;40(2):359–364.

10. Barry VC, Dillon T. Occurrence of xylans in marine algae. Nature. 1940;146(3706):620–620. https://doi.org/10.1038/146620a0

11. Percival EGV, Chanda SK. The xylan of Rhodymenia palmate. Nature. 1950;166(4227):787–788. https://doi.org/10.1038/166787b0

12. Kaur A, Singh A, Patra AK, Mahajan R. Costeffective scouring of flax fibers using cellulase-free xylano-pectinolytic synergism from a bacterial isolate. Journal of Cleaner Production. 2016;131:107–111. https://doi.org/10.1016/j.jclepro.2016.05.069

13. Akin DE. Plant cell wall aromatics: influence on degradation of biomass. Biofuels, Bioproducts and Biorefining. 2008;2:288–303. https://doi.org/10.1002/bbb.76

14. Kaur A, Mahajan R, Singh A, Garg G, Sharma J. A novel and cost effective methodology for qualitative screening of alkalothermophilic cellulase free xylano-pectinolytic microorganisms using agricultural wastes. World Journal of Microbiology and Biotechnology. 2011;27:459–463. https://doi.org/10.1007/s11274-010-0457-9

15. Subramaniyan S, Prema P. Biotechnology of microbial xylanases: Enzymology, molecular biology and application. Critical Reviews in Biotechnology. 2002;22(1):33–64. https://doi.org/10.1080/07388550290789450

16. Velázquez E, de Miguel T, Poza M, Rivas R, Rosselló-Mora R, Villa TG. Paenibacillus favisporus sp. nov., a xylanolytic bacterium isolated from cow faeces. International Journal of Systematic and Evolutionary Microbiology. 2004;54(1):59–64. https://doi.org/10.1099/ijs.0.02709-0

17. Rivas R, Mateos PF, Martínez-Molina E, Velázquez E. Paenibacillus phyllosphaerae sp. nov., a xylanolytic bacterium isolated from the phyllosphere of Phoenix dactylifera. International Journal of Systematic and Evolutionary Microbiology. 2005; 55(2):743–746. https://doi.org/10.1099/ijs.0.63323-0

18. Sánchez MM, Fritze D, Blanco A, Spröer C, Tindall BJ, Schumann P, et al. Paenibacillus barcinonensis sp. nov., a xylanase-producing bacterium isolated from a rice field in the Ebro River delta. International Journal of Systematic and Evolutionary Microbiology. 2005;55(2):935–939. https://doi.org/10.1099/ijs.0.63383-0

19. Ten LN, Baek SH, Im WT, Lee M, Oh HW, Lee ST. Paenibacillus panacisoli sp. nov., a xylanolytic bacterium isolated from soil in a ginseng field in South Korea. International Journal of Systematic and Evolutionary Microbiology. 2006;56(11):2677–2681. http://dx.doi.org/10.1099/ijs.0.64405-0

20. Rafikova GF, Loginova EV, Melentev AI, Loginov ON. Biological basis of microbial fodder additive. Patent RF, no. 2 662 931, 2018. (In Russian)

21. Li X, Yang SH, Yu XC, Jin ZX, Li WD, Li L, et al. Construction of transgenic Bacillus mucilaginosus strain with improved phytase secretion. Journal of Applied Microbiology. 2005;99(4):878–884. https://doi.org/10.1111/j.1365-2672.2005.02683.x

22. Von Schoultz S, AB BLN-WOODS LTD. Method for extracting biomass. U.S. Patent Application. 14/413,409. 2015. https://doi.org/10.1080/00031305.1999.10474445

23. Czitrom V. One-factor-at-a-time versus designed experiments. The American Statistician. 1999;53(2):126–131.

24. Maier RM. Environmental microbiology (second edition). Chapter 3. Bacterial growth. Elsevier Inc. 2009. Р. 37–54. https://doi.org/10.1016/B978-0-12-370519-8.00003-1

25. Bailey M, Biely P, Poutanen K. Interlaboratory testing of methods for assay of xylanase activity. Journal of Biotechnology. 1992;23(3):257–270. https://doi.org/10.1016/0168-1656(92)90074-J

26. Morozova YuA., Skvortsov EV, Alimova FK, Kanarsky AV. Biosynthesis of xylanases and cellulases by fungi Trichoderma on the post-alcohol bard. Vestnik Kazanskogo tekhnologicheskogo universiteta = Bulletin of Kazan Technological University. 2012;15(19):120–122. (In Russian)

27. Dias FE, Okrend H, Dondero NC. Growthpromoting activity of spent sulfite liquor for Sphaerotilus natans growing in a continuous-flow apparatus. Applied Microbiology. 1968;16(2):276–278. https://doi.org/10.1128/aem.16.2.276-278.1968

28. Patrauchan MA, Sarkisova S, Sauer K, Franklin MJ. Calcium influences cellular and extracellular product formation during biofilm-associated growth of a marine Pseudoalteromonas sp. Microbiology. 2005;151(9):2885–2897. https://doi.org/10.1099/mic.0.28041-0

29. Lamed R, Setiter E, Bayer EA. Characterization of a cellulose-binding, cellulase-containing complex in Clostridium thermocellum. Journal of Bacteriology. 1983;156(2):828–836. https://doi.org/10.1128/JB.156.2.828-836.1983

30. Grepinet O, Chebrou MC, Beguin P. Nucleotide sequence and deletion analysis of the xylanase gene (xynZ) of Clostridium thermocellum. Journal of Bacteriology. 1988;170(10):4582–4588. https://doi.org/10.1128/jb.170.10.4582-4588.1988

31. Ratanakhanokchai K, Kyu KL, Tanticharoen M. Purification and properties of a xylan-binding endoxylanase from alkaliphilic Bacillus sp. strain K-1. Applied and Environmental Microbiology. 1999; 65(2):694–697. https://doi.org/10.1128/AEM.65.2.694-697.1999

32. Howieson JG, Robson AD, Abbott LK. Calcium modifies pH effects on acid-tolerant and acidsensitive strains of Rhizobium meliloti. Australian Journal of Agricultural Research. 1992;43(3):765–772. https://doi.org/10.1071/AR9920765

33. Haltrich D, Nidetzky B, Kulbe KD, Steiner W, Župančič S. Production of fungal xylanases. Bioresource Technology. 1996;58(2):137–161. https://doi.org/10.1016/S0960-8524(96)00094-6

34. Kulkarni N, Shendye A, Rao M. Molecular and biotechnological aspects of xylanases. FEMS Microbiology Reviews. 1999;23(4):411–456. https://doi.org/10.1111/j.1574-6976.1999.tb00407.x

35. Ko C-H, Lin Z-P, Tu J, Tsai C-H, Liu C-C, Chen H-T, et al. Xylanase production by Paenibacillus campinasensis BL11 and its pretreatment of hardwood kraft pulp bleaching. International Biodeterioration & Biodegradation. 2010;64(1):13–19. https://doi.org/10.1016/j.ibiod.2009.10.001