Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Characterization of Aspergillus sojae Isolated from Meju, Korean Traditional Fermented Soybean Brick

  • Kim, Kyung Min (Department of Bio and Fermentation Convergence Technology, BK21 PLUS Project, Kookmin University) ;
  • Lim, Jaeho (Department of Bio and Fermentation Convergence Technology, BK21 PLUS Project, Kookmin University) ;
  • Lee, Jae Jung (Sempio Fermentation Research Center, Sempio Foods Company) ;
  • Hurh, Byung-Serk (Sempio Fermentation Research Center, Sempio Foods Company) ;
  • Lee, Inhyung (Department of Bio and Fermentation Convergence Technology, BK21 PLUS Project, Kookmin University)
  • Received : 2016.10.07
  • Accepted : 2016.11.21
  • Published : 2017.02.28
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Abstract

Initially, we screened 18 Aspergillus sojae-like strains from Aspergillus spp. isolated from meju (Korean traditional fermented soybean brick) according to their morphological characteristics. Because members of Aspergillus section Flavi are often incorrectly identified because of their phylogenetic similarity, we re-identified these strains at the morphological and molecular genetic levels. Fourteen strains were finally identified as A. sojae. The isolates produced protease and ${\alpha}-amylase$ with ranges of 2.66-10.64 and 21.53-106.73 unit/g-initial dry substrate (U/g-IDS), respectively, which were equivalent to those of the koji (starter mold) strains employed to produce Japanese soy sauce. Among the isolates and Japanese koji strains, strains SMF 127 and SMF 131 had the highest leucine aminopeptidase (LAP) activities at 6.00 and 6.06 U/g-IDS, respectively. LAP plays an important role in flavor development because of the production of low-molecular-weight peptides that affect the taste and decrease bitterness. SMF 127 and SMF 131 appeared to be non-aflatoxigenic because of a termination point mutation in aflR and the lack of the polyketide synthase gene found in other A. sojae strains. In addition, SMF 127 and SMF 131 were not cyclopiazonic acid (CPA) producers because of the deletion of maoA, dmaT, and pks/nrps, which are involved in CPA biosynthesis. Therefore, A. sojae strains such as SMF 127 and SMF 131, which have high protease and LAP activities and are free of safety issues, can be considered good starters for soybean fermentations, such as in the production of the Korean fermented soybean products meju, doenjang, and ganjang.

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References

  1. Sato A, Oshima K, Noguchi H, Ogawa M, Takahashi T, Oguma T, et al. 2011. Draft genome sequencing and comparative analysis of Aspergillus sojae NBRC4239. DNA Res. 18: 165-176. https://doi.org/10.1093/dnares/dsr009
  2. Kitamoto K. 2002. Molecular biology of the koji molds. Adv. Appl. Microbiol. 51: 129-154.
  3. Nampoothiri KM, Nagy V, Kovacs K, Szakacs G, Pandey A. 2005. L-Leucine aminopeptidase production by filamentous Aspergillus fungi. Lett. Appl. Microbiol. 41: 498-504. https://doi.org/10.1111/j.1472-765X.2005.01789.x
  4. Toldra F, Aristoy M-C, Flores M. 2000. Contribution of muscle aminopeptidases to flavor development in dry-cured ham. Food Res. Int. 33: 181-185. https://doi.org/10.1016/S0963-9969(00)00032-6
  5. Kim D-H, Kim S-H, Kwon S-W, Lee J-K, Hong S-B. 2013. Mycoflora of soybeans used for meju fermentation. Mycobiology 41: 100-107. https://doi.org/10.5941/MYCO.2013.41.2.100
  6. Buyukkileci AO, Tari C, Fernandez-Lahore M. 2011. Enhanced production of exo-polygalacturonase from agrobased products by Aspergillus sojae. Bioresources 6: 3452-3468.
  7. Chang P-K. 2004. Lack of interaction between AFLR and AFLJ contributes to nonaflatoxigenicity of Aspergillus sojae. J. Biotechnol. 107: 245-253. https://doi.org/10.1016/j.jbiotec.2003.10.012
  8. Gurkok S, Cekmecelioglu D, Ogel ZB. 2011. Optimization of culture conditions for Aspergillus sojae expressing an Aspergillus fumigatus ${\alpha}$-galactosidase. Bioresour. Technol. 102: 4925-4929. https://doi.org/10.1016/j.biortech.2011.01.036
  9. Heerd D, Yegin S, Tari C, Fernandez-Lahore M. 2012. Pectinase enzyme-complex production by Aspergillus spp. in solid-state fermentation: a comparative study. Food Bioprod. Process. 90: 102-110. https://doi.org/10.1016/j.fbp.2011.08.003
  10. Lin C-H, Wei Y-T, Chou C-C. 2006. Enhanced antioxidative activity of soybean koji prepared with various filamentous fungi. Food Microbiol. 23: 628-633. https://doi.org/10.1016/j.fm.2005.12.004
  11. Oncu S, Tari C, Unluturk S. 2007. Effect of various process parameters on morphology, rheology, and polygalacturonase production by Aspergillus sojae in a batch bioreactor. Biotechnol. Prog. 23: 836-845. https://doi.org/10.1002/bp070079c
  12. Chang P-K, Matsushima K, Takahashi T, Yu J, Abe K, Bhatnagar D, et al. 2007. Understanding nonaflatoxigenicity of Aspergillus sojae: a windfall of aflatoxin biosynthesis research. Appl. Microbiol. Biotechnol. 76: 977-984. https://doi.org/10.1007/s00253-007-1116-4
  13. Wicklow DT. 1984. Conidium germination rate in wild and domesticated yellow-green aspergilli. Appl. Environ. Microbiol. 47: 299-300.
  14. Chang H-Y, Lee Y-B, Bae H-A, Huh J-Y, Nam S-H, Sohn HS, et al. 2011. Purification and characterisation of Aspergillus sojae naringinase: the production of prunin exhibiting markedly enhanced solubility with in vitro inhibition of HMG-CoA reductase. Food Chem. 124: 234-241. https://doi.org/10.1016/j.foodchem.2010.06.024
  15. Murakami H, Hayashi K, Ushijima S. 1982. Useful key characters separating three Aspergillus taxa: A. sojae, A. parasiticus, and A. toxicarius. J. Gen. Appl. Microbiol. 28: 55-60. https://doi.org/10.2323/jgam.28.55
  16. Klich M, Mullaney E. 1989. Use of a bleomycin-containing medium to distinguish Aspergillus parasiticus from A. sojae. Mycologia 81: 159-160. https://doi.org/10.2307/3759464
  17. Glass NL, Donaldson GC. 1995. Development of primer sets designed for use with the PCR to amplify conserved genes from filamentous ascomycetes. Appl. Environ. Microbiol. 61: 1323-1330.
  18. Hubka V, Kolarik M. 2012. ${\beta}$-Tubulin paralogue tubC is frequently misidentified as the benA gene in Aspergillus section Nigri taxonomy: primer specificity testing and taxonomic consequences. Persoonia 29: 1. https://doi.org/10.3767/003158512X658123
  19. Peterson SW. 2008. Phylogenetic analysis of Aspergillus species using DNA sequences from four loci. Mycologia 100: 205-226. https://doi.org/10.1080/15572536.2008.11832477
  20. Saitou N, Nei M. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol. 4: 406-425.
  21. Godet M, Munaut F. 2010. Molecular strategy for identification in Aspergillus section Flavi. FEMS Microbiol. Lett. 304: 157-168. https://doi.org/10.1111/j.1574-6968.2009.01890.x
  22. Yuan GF, Liu CS, Chen CC. 1995. Differentiation of Aspergillus parasiticus from Aspergillus sojae by random amplification of polymorphic DNA. Appl. Environ. Microbiol. 61: 2384-2387.
  23. Miller GL. 1959. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem. 31: 426-428. https://doi.org/10.1021/ac60147a030
  24. Sahnoun M, Kriaa M, Elgharbi F, Ayadi D-Z, Bejar S, Kammoun R. 2015. Aspergillus oryzae S2 alpha-amylase production under solid state fermentation: optimization of culture conditions. Int. J. Biol. Macromol. 75: 73-80. https://doi.org/10.1016/j.ijbiomac.2015.01.026
  25. Kum S-J, Yang S-O, Lee SM, Chang P-S, Choi YH, Lee JJ, et al. 2015. Effects of Aspergillus species inoculation and their enzymatic activities on the formation of volatile components in fermented soybean paste (doenjang). J. Agric. Food Chem. 63: 1401-1418. https://doi.org/10.1021/jf5056002
  26. Cupp-Enyard C. 2008. Sigma's non-specific protease activity assay - casein as a substrate. J. Vis. Exp. 19: e899.
  27. Tan PST, Konings WN. 1990. Purification and characterization of an aminopeptidase from Lactococcus lactis subsp. cremoris Wg2. Appl. Environ. Microbiol. 56: 526-532.
  28. Lee JH, Jo EH, Hong EJ, Kim KM, Lee I. 2014. Safety evaluation of filamentous fungi isolated from industrial doenjang koji. J. Microbiol. Biotechnol. 24: 1397-1404. https://doi.org/10.4014/jmb.1403.03007
  29. Jorgensen TR. 2007. Identification and toxigenic potential of the industrially important fungi, Aspergillus oryzae and Aspergillus sojae. J. Food Prot. 70: 2916-2934. https://doi.org/10.4315/0362-028X-70.12.2916
  30. Murakami H. 1971. Classification of the koji mold. J. Gen. Appl. Microbiol. 17: 281-309. https://doi.org/10.2323/jgam.17.281
  31. Laforgue R, Guerin L, Pernelle JJ, Monnet C, Dupont J, Bouix M. 2009. Evaluation of PCR-DGGE methodology to monitor fungal communities on grapes. J. Appl. Microbiol. 107: 1208-1218. https://doi.org/10.1111/j.1365-2672.2009.04309.x
  32. Lee C-Z, Liou G-Y, Yuan G-F. 2006. Comparison of the aflR gene sequences of strains in Aspergillus section Flavi. Microbiology 152: 161-170. https://doi.org/10.1099/mic.0.27618-0
  33. Watson AJ, Fuller LJ, Jeenes DJ, Archer DB. 1999. Homologs of aflatoxin biosynthesis genes and sequence of aflR in Aspergillus oryzae and Aspergillus sojae. Appl. Environ. Microbiol. 65: 307-310.
  34. Chang P-K, Horn BW, Dorner JW. 2009. Clustered genes involved in cyclopiazonic acid production are next to the aflatoxin biosynthesis gene cluster in Aspergillus flavus. Fungal Genet. Biol. 46: 176-182. https://doi.org/10.1016/j.fgb.2008.11.002
  35. Sardjono, Zhu Y, Knol W. 1998. Comparison of fermentation profiles between lupine and soybean by Aspergillus oryzae and Aspergillus sojae in solid-state culture systems. J. Agric. Food Chem. 46: 3376-3380. https://doi.org/10.1021/jf980221c
  36. Felsenstein J. 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39: 783-791. https://doi.org/10.1111/j.1558-5646.1985.tb00420.x
  37. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. 2013. MEGA6: molecular evolutionary genetics analysis version 6.0. Mol. Biol. Evol. 30: 2725-2729. https://doi.org/10.1093/molbev/mst197
  38. Tominaga M, Lee Y-H, Hayashi R, Suzuki Y, Yamada O, Sakamoto K, et al. 2006. Molecular analysis of an inactive aflatoxin biosynthesis gene cluster in Aspergillus oryzae RIB strains. Appl. Environ. Microbiol. 72: 484-490. https://doi.org/10.1128/AEM.72.1.484-490.2006
  39. Chang P-K, Bhatnagar D, Cleveland TE, Bennett JW. 1995. Sequence variability in homologs of the aflatoxin pathway gene aflR distinguishes species in Aspergillus section Flavi. Appl. Environ. Microbiol. 61: 40-43.

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