Subjects: Biology >> Bioengineering submitted time 2019-04-21 Cooperative journals: 《中国生物工程杂志》
Abstract： 6-Hydroxynicotinic acid (6HNA) 3-monooxygenase (NicC) is the key enzyme for nicotinic degradation in Pseudomonas putida KT2440. NicC can catalyze the hydroxylation of pyridine ring to promote the ring cleavage reaction of pyridine ring. The expression level of NicC was enhanced by replace the rare codon in the N-terminal of NicC, and then the His-tagged NicC was purified to homogeneity. The optimal temperature reaction range of NicC is from 30℃ to 40℃, and the optimal reaction pH is 8.0. The Cd 2+ ion could significantly inhibit the activity of NicC. The apparent Km and Vmax values of the purified NicC for 6HNA were 51.8 μM and 14.1U/mg, respectively, and those for NADH were 15.0 μM and 10.79 U/mg, respectively. According to the HPLC and LC-MS analysis, NicC could catalyzes 6HNA to form 2,5-DHP and formic acid, and it could also transform 4-hydroxybenzoic acid to form hydroquinone. Isotope labeling experiments proved that the oxygen atom incorporated into 2,5-DHP is from dioxygen. The study will provide useful information for the microbial degradation of pyridinic compounds.
Subjects: Biology >> Bioengineering submitted time 2017-07-24 Cooperative journals: 《中国生物工程杂志》
Abstract：摘 要：从辽河口石油污染土壤中筛选到一株能够以2-羟基吡啶作为唯一碳源、氮源和能源进行生长的菌株2PR，基于形态学观察、16S rRNA基因序列分析鉴定菌株2PR属于节杆菌属（Arthrobacter）。菌株2PR生长和降解2-羟基吡啶的最适条件是30℃，pH为7.0。当2-羟基吡啶初始浓度为6.0 mg/mL时，120 h菌株2PR对2-羟基吡啶的降解效率为94.48%，初始2-羟基吡啶浓度为8.0 mg/mL时，156 h的降解效率为89.21%。对2-羟基吡啶降解动力学过程进行模拟，结果显示菌株2PR生长和降解过程符合logisitic模型，该模型为环境中2-羟基吡啶的生物降解提供了理论参考。休止细胞反应和中间代谢产物检测表明，菌株2PR在降解2-羟基吡啶的过程中生成了蓝色化合物4,5,4′,5′-tetrahydroxy-3,3′-diazadiphenoquinone-(2,2′)。推测该菌株降解2-羟基吡啶的途径可能是首先由双加氧酶催化生成2,3,6-三羟基吡啶，后者会自发形成蓝色中间代谢产物，2,3,6-三羟基吡啶发生开环反应，最终被完全降解。菌株2PR是已报道菌株中2-羟基吡啶耐受能力和降解能力最强的菌株，在污染物生物修复方面具有广阔的应用前景。
Subjects: Biology >> Bioengineering submitted time 2017-06-01 Cooperative journals: 《中国生物工程杂志》
Abstract： Abstract: A novel strain, which could use 2-hydroxypyridine (2HP) as the sole source of carbon, nitrogen, and energy, was isolated from petroleum-contaminated soil at the Liaohe estuarine wetland. Strain 2PR was identified as Arthrobacter based on the morphology and 16S rRNA gene sequence. The optimum growth and degradation condition upon 2PR is 30℃ and pH 7.0, respectively. Under this condition, 2HP degradation rate were 97.34%, 94.95%, 94.48% and 89.21% with 2, 4, 6 and 8 mg/mL initial concentration of 2HP at 42, 96, 120 and 156 h, respectively. Strain growth and 2HP degradation kinetics studies indicated that the strain followed Logisitic model, which could provide a theoretical and technical reference for the biodegradation of 2HP. The color of strain 2PR culture upon 2HP-MSN changed from colorless to blue, and then turned to brown. The blue pigment, which was observed at the culture of strain 2PR, was identified as 4,5,4′,5′-tetrahydroxy-3,3′-diazadiphenoquinone-(2,2′) by high performance liquid chromatography (HPLC) and high-performance liquid chromatography-mass spectrometry (LC-MS) analysis. The LC-MS signal with m/z=249.1 was observed in resting cells reaction sample with 2HP as the substrate. The degradation of 2HP might be achieved by a dioxygenase to produce 2,3,6-trihydroxypyridine, which could transformed to the blue pigment spontaneously, and then 2,3,6-trihydroxypyridine was converted with an pyridine-ring cleavage reaction. Among all the reported strains, strain 2PR has the strongest tolerance and the highest 2HP degradation efficiency at present. The strain has a promising application potential for 2HP waste treatment.
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