Properties of a Poly(3-hydroxybutyrate) Depolymerase from Penicillium funiculosum
Properties of a Poly(3-hydroxybutyrate) Depolymerase from Penicillium funiculosum
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绳状青霉聚（3-羟基丁酸酯）解聚酶的特性
DOI：
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发表时间：
2000
期刊：
影响因子：
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通讯作者：
K. Kasuya
中科院分区：
文献类型：
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作者：
Sakie Miyazaki;Kazuhei Takahashi;M. Shiraki;Terumi Saito;Y. Tezuka;K. Kasuya
用苯基托菌灵从绳状青霉（Penicillium funculosum，IFO 6345）中纯化了一种聚3-羟基丁酸酯（poly（3-hydroxybutyrate，PHB））解聚酶，并将其性质与其它几种PHB解聚酶进行了比较。通过十二烷基硫酸钠（SDS）聚丙烯酰胺凝胶电泳，纯化的酶的分子量估计约为33 kDa。最适pH和pI分别为6.5和6.5。纯化的蛋白显示出对Con A-Sepharose的亲和力，表明其为糖蛋白。二异丙基氟磷酸和二硫苏糖醇完全抑制解聚酶活性。纯化的酶的N-末端氨基酸序列为TALPAFNVNPNSVSVSGLSSGGYMAAQL，其含有“脂肪酶盒”序列。该酶是丝氨酸酯酶中的一种胞外PHB解聚酶。纯化的酶表现出相对较强的水解活性对3-羟基丁酸酯低聚物相比，其聚羟基丁酸酯降解活性。聚羟基丁酸结合实验表明，绳状青霉解聚酶具有最弱的亲和力的所有解聚酶检查的聚羟基丁酸。
A poly(3-hydroxybutyrate) (PHB) depolymerase was purified from a fungus, Penicillium funiculosum (IFO6345), with phenyl-Toyopearl and its properties were compared with those of other PHB depolymerases. The molecular mass of the purified enzyme was estimated at about 33 kDa by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis. The pH optimum and pI were 6.5 and 6.5, respectively. The purified protein showed affinity to Con A-Sepharose, indicating that it is a glycoprotein. Diisopropylfluorophosphate and dithiothreitol inhibited the depolymerase activity completely. The N-terminal amino acid sequence of the purified enzyme was TALPAFNVNPNSVSVSGLSSGGYMAAQL, which contained a “lipase box” sequence. This purified enzyme is one of the extracellular PHB depolymerase which belong to serine esterase. The purified enzyme showed relatively strong hydrolytic activity against 3-hydroxybutyrate oligomers compared with its PHB-degrading activity. PHB-binding experiments showed that P. funiculosum depolymerase has the weakest affinity for PHB of all the depolymerases examined.