基于宏基因组学的海带降解菌群微生物多样性及其褐藻多糖降解酶系分析
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摘要
为系统探讨一组高效海带降解菌群的微生物多样性及其褐藻多糖降解酶系,本实验利用高通量测序技术对该菌群进行组学分析。结果表明,共获得36552330条reads,预测得到105145条基因序列,比对显示此高效海带降解菌群中有75.41%的厚壁菌门(Firmicutes)细菌,优势菌属包括Lachnoclostridium、芽孢杆菌属(Bacillus)、类芽孢杆菌属(Paenibacillus)、解硫胺素芽孢杆菌属(Aneurinibacillus)、短芽孢杆菌属(Brevibacillus)。数据显示有4526个基因编码蛋白质具有海带多糖降解相关酶活性,其中糖苷水解酶(GH)基因数量最多(1520),碳水化合物结合模块(CBM)(764)、碳水化合物酯酶(CE)基因(706)和糖基转移酶(GT)(693)数量次之,多糖裂合酶(PL)基因(107)和辅助氧化还原酶(AA)基因(126)数量较少。在不同的GH基因家族,归属于GH109、GH13、GH18、GH43家族的基因较多。此外还发现了少量的纤维小体组分蛋白基因。本研究发现海带降解菌群中有丰富的褐藻胶裂解酶,纤维素酶,果胶酶,淀粉酶酶系产酶菌,为进一步开发人工高效褐藻降解菌群及褐藻多糖降解酶提供参考。
In order to investigate systematically the diversity and brown algal polysaccharide degrading enzymes of Laminaria japonica degrading microbes,the metagenome of these microbes was sequenced with massively parallel sequencing technology in this study.A total of 36,552,330 reads were obtained and 105,145 genes were modeled.The results showed that the microbes in phylum Firmicutes accounted for 75.41%among the microbes digesting L.japonica.The dominant bacteria were from genera Lachnoclostridium,Bacillus,Paenibacillus,Aneurinibacillus and Brevibacillus.In total,4,526 genes were found to encode enzymes degrading L.japonica.The number of glycoside hydrolase(GH)genes was the largest(1,520),which was followed by that of carbohydrate binding module(CBM)genes(764),carbohydrate esterase(CE)genes(706)and glycosyl transferase(GT)genes(693).The polysaccharide lyase(PL)genes(107)and their auxiliary protein(AP)genes(126)were the two least.A large portion of GH genes belonged to GH109,GH13,GH18 and GH43 families.In addition,a small number of cellulosome component protein genes were found.We found also abundant alginate lyase,cellulase,pectinase and amylase producing bacteria,which provided basic data for further technological developments of highly efficient artificial degradation of L.japonicaand brown algal polysaccharide exploitation.
In order to investigate systematically the diversity and brown algal polysaccharide degrading enzymes of Laminaria japonica degrading microbes,the metagenome of these microbes was sequenced with massively parallel sequencing technology in this study.A total of 36,552,330 reads were obtained and 105,145 genes were modeled.The results showed that the microbes in phylum Firmicutes accounted for 75.41%among the microbes digesting L.japonica.The dominant bacteria were from genera Lachnoclostridium,Bacillus,Paenibacillus,Aneurinibacillus and Brevibacillus.In total,4,526 genes were found to encode enzymes degrading L.japonica.The number of glycoside hydrolase(GH)genes was the largest(1,520),which was followed by that of carbohydrate binding module(CBM)genes(764),carbohydrate esterase(CE)genes(706)and glycosyl transferase(GT)genes(693).The polysaccharide lyase(PL)genes(107)and their auxiliary protein(AP)genes(126)were the two least.A large portion of GH genes belonged to GH109,GH13,GH18 and GH43 families.In addition,a small number of cellulosome component protein genes were found.We found also abundant alginate lyase,cellulase,pectinase and amylase producing bacteria,which provided basic data for further technological developments of highly efficient artificial degradation of L.japonicaand brown algal polysaccharide exploitation.
引文
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[18]孙丕海,钱坤,李晓丽,等.基于高通量测序分析海带表面细菌群落结构[J].大连:大连海洋大学学报,2017,32(1):7-12.
[19]侯士昌,温少红,唐志红,等.一株高效褐藻酸降解菌的筛选、鉴定及其发酵条件的优化[J].北京:海洋科学,2014,38(7):20-26.
[20]吴丽云,倪辉,李鹤宾,等.微泡菌属ALW5褐藻胶裂解酶的酶学性质及酶解产物的抗氧化活性[J].北京:中国食品学报,2016,16(6):79-87.
[21] Zhu Y,Wu L,Chen Y,et al.Characterization of an extracellular biofunctional alginate lyase from marine Microbulbifer sp.ALW1and antioxidant activity of enzymatic hydrolysates[J].Germany:Microbiological Research,2016,182:49.
[22] Mohapatra B R.Kinetic and thermodynamic properties of alginate lyase and cellulase co-produced by Exiguobacterium species Alg-S5.[J].Netherlands:International Journal of Biological Macromolecules,2017,98:103-110.
[23]张小梅,李单单,王禄山,等.纤维素酶家族及其催化结构域分子改造的新进展[J].北京:生物工程学报,2013,29(4):422-433.
[24] Patel D D,Patel A K,Parmar N R,et al.Microbial and Carbohydrate Active Enzyme profile of buffalo rumen metagenome and their alteration in response to variation in the diet[J].Netherlands:Gene,2014,545(1):88.
[25] Zhang H,Li J,Wang J,et al.Determinants for the improved thermostability of a mesophilic family 11xylanase predicted by computational methods[J].England:Biotechnology for Biofuels,2014,7(1):3.
[26] Yadav S,Yadav P K,Yadav D,et al.Pectin lyase:a review.[J].England:Process Biochemistry,2009,44(1):1-10.
[27]周燕霞.褐藻胶裂解酶分泌菌株的分离鉴定及Tamlana holothuriorum s12~T中褐藻胶裂解酶的研究[D].济南:山东大学,2016,4-5.
[28]张慧敏,夏海磊,黄强,等.海子水牛瘤胃微生物的宏基因组学分析[J].北京:动物营养学报,2017,29(11).
[29] Hess M,Sczyrba A,Egan R,et al.Metagenomic discovery of biomass-degrading genes and genomes from cow rumen[J].Washington:Science,2011,331(6016):463-467.
[2]郭肖青.海藻纤维的制备及结构与性能研究[D].青岛:青岛大学,2007.
[3]沙源,夏延致,王兵兵,等.非水溶性天然色素原液着色海藻纤维的制备及其性能[J].北京:纺织学报,2017,38(9):1-7.
[4] Chiu K W,Fung A Y.The cardiovascular effects of green beans(Phaseolus aureus),common rue(Ruta graveolens),and kelp(Laminaria japonica)in rats[J].USA:Phytotherapy Research,2015,11(3):203-206.
[5] Zhao C,Yang C,Liu B,et al.Bioactive compounds from marine macroalgae and their hypoglycemic benefits[J].Trends in Food Science&Technology,USA:2017,72.
[6] Zhao X,Guo F,Hu J,et al.Antithrombotic activity of oral administered low molecular weight fucoidan fromLaminaria japonica[J].England:Thrombosis Research,2016,144:46-52.
[7] Zhu Q,Chen J,Li Q,et al.Antitumor activity of polysaccharide fromLaminaria japonica,on mice bearing H22liver cancer[J].Netherlands:International Journal of Biological Macromolecules,2016,92:156-158.
[8] Zha X Q,Lu C Q,Cui S H,et al.Structural identification and immunostimulating activity of a Laminaria japonica,polysaccharide[J].Netherlands:International Journal of Biological Macromolecules,2015,78:429-438.
[9] Fang Q,Wang J F,Zha X Q,et al.Immunomodulatory activity on macrophage of a purified polysaccharide extracted fromLaminaria japonica[J].England:Carbohydrate Polymers,2015,134:66-73.
[10] Cao Y G,Hao Y,Li Z H,et al.Antiviral activity of polysaccharide extract fromLaminaria japonica,against respiratory syncytial virus[J].Paris:Biomedicine&Pharmacotherapy,2016,84(86):1705-1710.
[11]陈朋,朱玥明,韩文佳,等.产褐藻胶裂解酶菌种的筛选、鉴定及发酵条件优化[J].北京:食品科学,2015,36(15):105-111.
[12]王熙涛,徐永平,金礼吉,等.具有海带褐藻胶降解能力的刺参有益菌筛选及降解条件优化[J].北京:中国饲料,2014(4):27-31.
[13]李婷婷,陈倩,石萍,等.高效褐藻胶降解菌的筛选与产酶条件优化[J].北京:北京大学学报(自然科学版),2017,53(6):1115-1121.
[14]魏丹,窦文芳,李恒,等.高效降解褐藻胶新菌种的筛选、鉴定及产酶条件优化[J].北京:食品与发酵工业,2012,38(7):26-31.
[15]柏超.海带降解复合菌的选育及其功能研究[D].杭州:浙江大学,2012.
[16] Yin Y B,Mao X Z,Yang J C,et al.dbCAN:a web resource for automated carbohydrate-active enzyme annotation.[J].Oxford:Nucleic Acids Research,2012,40(Web Server issue):445-51.
[17]张彩凤,王婷婷,高小宽,等.宏基因组学技术及其应用概述[J].上海:生物学教学,2013,38(3):7-8.
[18]孙丕海,钱坤,李晓丽,等.基于高通量测序分析海带表面细菌群落结构[J].大连:大连海洋大学学报,2017,32(1):7-12.
[19]侯士昌,温少红,唐志红,等.一株高效褐藻酸降解菌的筛选、鉴定及其发酵条件的优化[J].北京:海洋科学,2014,38(7):20-26.
[20]吴丽云,倪辉,李鹤宾,等.微泡菌属ALW5褐藻胶裂解酶的酶学性质及酶解产物的抗氧化活性[J].北京:中国食品学报,2016,16(6):79-87.
[21] Zhu Y,Wu L,Chen Y,et al.Characterization of an extracellular biofunctional alginate lyase from marine Microbulbifer sp.ALW1and antioxidant activity of enzymatic hydrolysates[J].Germany:Microbiological Research,2016,182:49.
[22] Mohapatra B R.Kinetic and thermodynamic properties of alginate lyase and cellulase co-produced by Exiguobacterium species Alg-S5.[J].Netherlands:International Journal of Biological Macromolecules,2017,98:103-110.
[23]张小梅,李单单,王禄山,等.纤维素酶家族及其催化结构域分子改造的新进展[J].北京:生物工程学报,2013,29(4):422-433.
[24] Patel D D,Patel A K,Parmar N R,et al.Microbial and Carbohydrate Active Enzyme profile of buffalo rumen metagenome and their alteration in response to variation in the diet[J].Netherlands:Gene,2014,545(1):88.
[25] Zhang H,Li J,Wang J,et al.Determinants for the improved thermostability of a mesophilic family 11xylanase predicted by computational methods[J].England:Biotechnology for Biofuels,2014,7(1):3.
[26] Yadav S,Yadav P K,Yadav D,et al.Pectin lyase:a review.[J].England:Process Biochemistry,2009,44(1):1-10.
[27]周燕霞.褐藻胶裂解酶分泌菌株的分离鉴定及Tamlana holothuriorum s12~T中褐藻胶裂解酶的研究[D].济南:山东大学,2016,4-5.
[28]张慧敏,夏海磊,黄强,等.海子水牛瘤胃微生物的宏基因组学分析[J].北京:动物营养学报,2017,29(11).
[29] Hess M,Sczyrba A,Egan R,et al.Metagenomic discovery of biomass-degrading genes and genomes from cow rumen[J].Washington:Science,2011,331(6016):463-467.