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Identification and structural analysis of novel laccase genes in Flammulina elastica genome

Flammulina elastica의 유전체 정보기반 신규 laccase 유전자 동정 및 구조 분석

  • Yu, Hye-Won (Department of Medicinal Biosciences, Konkuk University) ;
  • Park, Young-Jin (Department of Medicinal Biosciences, Konkuk University)
  • 유혜원 (건국대학교 글로컬캠퍼스 의료생명대학 바이오의약학과) ;
  • 박영진 (건국대학교 글로컬캠퍼스 의료생명대학 바이오의약학과)
  • Received : 2021.02.03
  • Accepted : 2021.03.02
  • Published : 2021.03.31

Abstract

The genome sequence of various Flammulina species has recently been reported, thereby revealing a diverse genetic repertoire. In this study, we identified laccase genes and analyzed their structural characteristics in Flammulina elastica (F. elastica) genome. Through genome analysis and bioinformatics approaches, three laccase genes (Fe-lac1, -lac2, and -lac3) were identified, ranging from 1,548 to 1,602 bp in length. These genes contained a copper ion-binding region with ten histidine residues and one cysteine residue and a disulfide bond-forming region with four cysteine residues. Full-length cDNA sequencing analysis revealed that laccase genes contain 12 to 16 introns and signal peptides between 17 and 22 bp in the N-terminus. Structural characterization of the laccase genes identified in this study should help in better understanding the biomass decomposition of F. elastica.

최근 Flammulina 종들에 대한 유전체 염기서열 분석결과가 보고되었고, 그로 인해 다양한 유전자 정보가 밝혀지고 있다. 본 연구에서는 Flammulina elastica 전체 유전체 서열의 laccase 유전자를 동정하고 구조적 특징 분석을 수행하고자 하였다. 유전체 분석 및 생물정보분석을 통하여 F. elastica 유전체 내 3개의 laccase 유전자(Fe-lac1, Fe-lac2, Fe-lac3)를 확인하였고, 이들 유전자 내에는 10개의 히스티딘 잔기와 1개의 시스테인 잔기를 가지는 구리 이온 결합 영역과 4개의 시스테인 잔기를 가지는 이황화결합 형성 부위가 존재하는 것을 확인하였다. 1,548~1,602 bp의 laccase 유전자에 대한 전장 cDNA 염기서열 분석을 통하여 12~16개의 인트론이 존재하는 것을 확인되었으며, N-말단으로부터 17~22 bp의 사이에 신호펩타이드가 존재하는 것이 확인되었다. 본 연구를 통하여 F. elastica의 laccase 유전자를 최초로 동정하여 구조적 특징을 분석하였고, 이러한 결과는 F. elastica의 바이오매스 분해에 대한 이해를 돕는데 활용될 것으로 사료된다.

Keywords

References

  1. Altschul SF, Erickson BW. 1985. Significance of nucleotide sequence alignments: a method for random sequence permutation that preserves dinucleotide and codon usage. Mol Biol Evol 2: 526-538.
  2. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. 1990. Basic local alignment search tool. J Mol Biol 215: 403-410. https://doi.org/10.1016/S0022-2836(05)80360-2
  3. Ballesteros M, Oliva JM, Negro MJ, Manzanares P, Ballesteros I. 2004. Ethanol from lignocellulosic materials by a simultaneous saccharification and fermentation process (SFS) with Kluyveromyces marxianus CECT 10875. Process Biochem 39: 1843-1848. https://doi.org/10.1016/j.procbio.2003.09.011
  4. Bento I, Carrondo MA, Lindley PF. 2006. Reduction of dioxygen by enzymes containing copper. J Biol Inorg Chem 11: 539-547. https://doi.org/10.1007/s00775-006-0114-9
  5. Bertrand T, Jolivalt C, Briozzo P, Caminade E, Joly N, Madzak C, Mougin C. 2002. Crystal structure of a four-copper laccase complexed with an arylamine: insights into substrate recognition and correlation with kinetics. Biochemistry 41: 7325-7333. https://doi.org/10.1021/bi0201318
  6. Bolger AM, Lohse M, Usadel B. 2014. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30: 2114-2120. https://doi.org/10.1093/bioinformatics/btu170
  7. Breathnach R, Benoist C, O'Hare K, Gannon F, Chambon P. 1978. Ovalbumin gene: evidence for a leader sequence in mRNA and DNA sequences at the exon-intron boundaries. Proc Natl Acad Sci USA 75: 4853-4857. https://doi.org/10.1073/pnas.75.10.4853
  8. Buchfink B, Xie C, Huson DH. 2015. Fast and sensitive protein alignment using DIAMOND. Nat Methods 12: 59-60. https://doi.org/10.1038/nmeth.3176
  9. Eggert C, Temp U, Dean JFD, Eriksson KEL. 1996. A fungal metabolite mediates degradation of non-phenolic lignin structures and synthetic lignin by laccase. FEBS Lett 391: 144-148. https://doi.org/10.1016/0014-5793(96)00719-3
  10. Eriksson KEL, Blanchette RA, Ander P. 1990. Morphological aspects of wood degradation by fungi and bacteria. In K. E. L. Eriksson, R. A. Blanchette & P. Ander. Microbial and Enzymatic Degradation of Wood and Wood Components, Springer-Verlag Berlin Heidelberg. Berlin, Heidelberg. 1-87.
  11. Finn RD, Coggill P, Eberhardt RY, Eddy SR, Mistry J, Mitchell AL, Potter SC, Punta M, Qureshi M, Sangrador-Vegas A, et al. 2016. The Pfam protein families database: towards a more sustainable future. Nucleic acids Res 44: D279-D285. https://doi.org/10.1093/nar/gkv1344
  12. Fitch WM. 1983. Random sequences. J Mol Biol 163: 171-176. https://doi.org/10.1016/0022-2836(83)90002-5
  13. Galhaup C, Goller S, Peterbauer CK, Strauss J, Haltrich D. 2002. Characterization of the major laccase isoenzyme from Trametes pubescens and regulation of its synthesis by metal ions. Microbiology 148: 2159-2169. https://doi.org/10.1099/00221287-148-7-2159
  14. Giardina P, Faraco V, Pezzella C, Piscitelli A, Vanhulle S, Sannia G. 2010. Laccases: a never-ending story. Cell Mol Life Sci 67: 369-385. https://doi.org/10.1007/s00018-009-0169-1
  15. Ha HC. 2012. Screening and production of lignocellulolytic enzymes secreted by the edible basidiomycete Pleurotus ostreatus. J Mushrooms 10: 74-82.
  16. Hakulinen N, Rouvinen J. 2015. Three-dimensional structures of laccases. Cell Mol Life Sci 72: 857-868. https://doi.org/10.1007/s00018-014-1827-5
  17. Hatamoto O, Sekine H, Nakano E, Abe K. 1999. Cloning and expression of a cDNA encoding the laccase from Schizophyllum commune. Biosci Biotechnol Biochem 63: 58-64. https://doi.org/10.1271/bbb.63.58
  18. Jain RG, Rusch SL, Kendall DA. 1994. Signal peptide cleavage regions. Functional limits on length and topological implications. J Biol Chem 269: 16305-16310. https://doi.org/10.1016/S0021-9258(17)34008-5
  19. Kim HI, Kwon OC, Kong WS, Lee CS, Park YJ. 2014. Genomewide identification and characterization of novel laccase genes in the white-rot fungus Flammulina velutipes. Mycobiology 42: 322-330. https://doi.org/10.5941/MYCO.2014.42.4.322
  20. Kwon JK, Moon HS, Kim JS, Kim SW, Hong SI. 1999. Fed-batch simultaneous saccharification and fermentation of waste paper to ethanol. Korean J Biotechnol Bioeng 14: 24-30.
  21. Lee SB, Lee JD. 2010. Effect of pretreatment process on cellulosic ethanol production using waste papers. J Korea Soc Waste Manag 27: 553-557.
  22. Lipman DJ, Wilbur WJ, Smith TF, Waterman MS. 1984. On the statistical significance of nucleic acid similarities. Nucleic Acids Res 12: 215-226. https://doi.org/10.1093/nar/12.1Part1.215
  23. Martone PT, Estevez JM, Lu F, Ruel K, Denny MW, Somerville C, Ralph J. 2009. Discovery of lignin in seaweed reveals convergent evolution of cell-wall architecture. Curr Biol 19: 169-175. https://doi.org/10.1016/j.cub.2008.12.031
  24. Mayer AM, Staples RC. 2002. Laccase: new functions for an old enzyme. Phytochemistry 60: 551-565. https://doi.org/10.1016/S0031-9422(02)00171-1
  25. Park YJ, Baek JH, Lee S, Kim C, Rhee H, Kim H, Seo JS, Park HR, Yoon DE, Nam JY, et al. 2014. Whole genome and global gene expression analyses of the model mushroom Flammulina velutipes reveal a high capacity for lignocellulose degradation. PLoS One 9: e93560. https://doi.org/10.1371/journal.pone.0093560
  26. Punta M, Coggill PC, Eberhardt RY, Mistry J, Tate J, Boursnell C, Pang N, Forslund K, Ceric G, Clements J, et al. 2012. The Pfam protein families database. Nucleic Acids Res 40: D290-D301. https://doi.org/10.1093/nar/gkr717
  27. Redhead SA, Petersen RH. 1999. New species, varieties and combinations in the genus Flammulina. Mycotaxon 71: 285-294.
  28. Rytioja J, Hilden K, Yuzon J, Hatakka A, de Vries RP, Makela MR. 2014. Plant-polysaccharide-degrading enzymes from basidiomycetes. Microbiol Mol Biol Rev 78: 614-649. https://doi.org/10.1128/MMBR.00035-14
  29. Scott CD, Davison BH, Scott TC, Woodward J, Dees C, Rothrock DS. 1994. An advanced bioprocessing concept for the conversion of waste paper to ethanol. Appl Biochem Biotechnol 45: 641-653. https://doi.org/10.1007/BF02941836
  30. Sista Kameshwar AK, Qin W. 2017. Comparative study of genome-wide plant biomass-degrading CAZymes in white rot, brown rot and soft rot fungi. Mycology 9: 93-105. https://doi.org/10.1080/21501203.2017.1419296
  31. Smale ST, Kadonaga JT. 2003. The RNA polymerase II core promoter. Annu Rev Biochem 72: 449-479. https://doi.org/10.1146/annurev.biochem.72.121801.161520
  32. Solomon EI, Sundaram UM, Machonkin TE. 1996. Multicopper oxidases and oxygenases. Chem Rev 96: 2563-2606. https://doi.org/10.1021/cr950046o
  33. Stanke M, Morgenstern B. 2005. AUGUSTUS: a web server for gene prediction in eukaryotes that allows user-defined constraints. Nucleic Acids Res 33: W465-W467. https://doi.org/10.1093/nar/gki458
  34. Strange RW, Reinhammer B, Murphy LM, Hasnain SS. 1995. Structural and spectroscopic studies of the copper site of stellacyanin. Biochemistry 34: 220-231. https://doi.org/10.1021/bi00001a026
  35. Thurston CF. 1994. The structure and function of fungal laccases. Microbiology 140: 19-26. https://doi.org/10.1099/13500872-140-1-19
  36. Weinzierl ROJ. 1999. Mechanisms of gene expression: structure, function and evolution of the basal transcriptional machinery. Imperial College Press, London. 1-424.
  37. Yaropolov AI, Skorobogat'ko OV, Vartanov SS, Varfolomeyev SD. 1994. Laccase properties, catalytic mechanism, and applicability. Appl Biochem Biotechnol 49: 257-280. https://doi.org/10.1007/BF02783061
  38. Zerbino DR, Birney E. 2008. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18: 821-829. https://doi.org/10.1101/gr.074492.107
  39. Zhu JY, Pan XJ. 2010. Woody biomass pretreatment for cellulosic ethanol production: technology and energy consumption evaluation. Bioresour Technol 101: 4992-5002. https://doi.org/10.1016/j.biortech.2009.11.007