一株海藻酸降解菌产酶条件的优化
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摘要
褐藻酸分解菌是褐藻酸裂解酶的重要来源,具有明确的应用价值。通过以海藻酸钠为唯一碳源培养基筛选的方法,从采集自长岛的腐烂海带组织中分离出1株高产褐藻酸裂解酶的新菌株B2,并对其进行了鉴定及培养条件优化。通过综合形态学鉴定和分子生物学鉴定确定该菌株属于副球属(Paracoccus)。利用响应面试验设计优化了该菌株产酶条件,得到最佳发酵产酶条件为发酵温度25℃,转速150 r/min,pH值为7.0,接种量8%,在此优化条件下,该菌株产褐藻酸裂解酶的酶活力为43.6 U/mol。
Alginate-degrading bacteria are important source of alginate lyase, which has certain application value. One new strain B2 with high-yield alginate lyase was isolated from the tissues of decaying kelp collected from Chang Island by a selection medium using sodium alginate as the sole carbon source. The strain B2 was identified and its fermentation conditions were optimized. The strain B2 was identified as Paracoccus by morphological identification and molecular biological identification. The enzyme-producing conditions were optimized by response surface experimental design and the optimal alginate lyase production condition was determined as fermentation temperature 25 ℃, rotational speed 150 r/min, pH value 7.0 and inoculum 8%. Under the optimal conditions, the alginate lyase activity of the strain was 43.6 U/mol.
Alginate-degrading bacteria are important source of alginate lyase, which has certain application value. One new strain B2 with high-yield alginate lyase was isolated from the tissues of decaying kelp collected from Chang Island by a selection medium using sodium alginate as the sole carbon source. The strain B2 was identified and its fermentation conditions were optimized. The strain B2 was identified as Paracoccus by morphological identification and molecular biological identification. The enzyme-producing conditions were optimized by response surface experimental design and the optimal alginate lyase production condition was determined as fermentation temperature 25 ℃, rotational speed 150 r/min, pH value 7.0 and inoculum 8%. Under the optimal conditions, the alginate lyase activity of the strain was 43.6 U/mol.
引文
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[2]周燕霞.褐藻胶裂解酶分泌菌株的分离鉴定及Tamlana holothuriorum s12T中褐藻胶裂解酶的研究[D].济南:山东大学,2016.
[3]GACESA P.Enzyme degradation of alginates[J].Int J Biochem,1992,24(4):545-552.
[4]SIM S J,BAIK K S,PARK S C,et al.Characterization of alginate lyase gene using a metagenomic library constructed from the gut microflora of abalone[J].J Ind Microbiol Biot,2012,39(4):585-593.
[5]宋鑫,秦送玉,朱军,等.产褐藻胶裂解酶海洋细菌的筛选和鉴定[J].基因组学与应用生物学,2016,35(4):907-912.
[6]韩峰.海洋弧菌QY103褐藻胶裂解酶的研究[D].青岛:中国海洋大学,2008.
[7]许超,黄桂媛,王巧贞,等.褐藻胶裂解酶的基因克隆及表达条件优化[J].中国酿造,2017,36(12):120-125.
[8]KIM D E,LEE E Y,KIM H S.Cloning and characterization of alginate lyase from a marine bacterium Streptomyces sp.ALG-5[J].Mar Biotechnol,2009,11(1):10-16.
[9]王迪.褐藻胶裂解菌株筛选及优化[D].大连:大连海洋大学,2017.
[10]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].北京大学学报:自然科学版,2017,53(6):1115-1121.
[11]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].海洋科学,2014,38(7):20-26.
[12]王林娜.海洋细菌Shewanella sp.Kz7的寡褐藻胶裂解酶研究[D].青岛:中国海洋大学,2014.
[13]BOX G E P,BEHNKEN D W.Simplex-sum designs:a class of second order rotatable designs derivable from those of first order[J].Ann Math Stat,1960,31(4):838-864.
[14]RAGONESE R,MACKA M,HUGHES J,et al.The use of the BoxBehnken experimental design in the optimisation and robustness testing of a capillary electrophoresis method for the analysis of ethambutol hydrochloride in a pharmaceutical formulation[J].J Pharmaceut Biomed,2002,27(6):995-1007.
[15]LEE K M,GILMORE D F.Statistical experimental design for bioprocess modeling and optimization analysis[J].Appl Biochem Biotech,2006,135(2):101-115.
[16]金德强,王超,翟乃明,等.响应面法优化修道院强烈艾尔啤酒酿造工艺的研究[J].中国酿造,2018,37(6):140-144.
[17]许超,熊亚茹,卢明倩,等.一株具有褐藻胶降解能力的海洋细菌的筛选鉴定及其多糖利用能力研究[J].生物技术通报,2017,33(4):198-204.
[18]林清菁,潘崇良,蒋霞敏,等.一株海藻多糖降解菌的分离与鉴定[J].微生物学通报,2014,41(11):2208-2215.
[19]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].食品科学,2015,36(15):105-111.
[20]SCHLEIFER K H.Microbial diversity:facts,problems and prospects[J].Syst Appl Microbiol,2004,27(1):3-9.
[21]李斌,何红波,李义兵.基于DNA序列LZ复杂性距离的系统进化树重构[J].高技术通讯,2006,16(5):506-510.
[22]DAVIS D H,DOUDOROFF M,STANIER R Y,et al.Proposal to reject the genus Hydrogenomonas:Taxonomic implications[J].Int J Syst Evol Mic,1969,19(4):375-390.