微紫青霉酸性木聚糖酶xynA基因的克隆与序列分析

发布时间：2018-03-18 22:45

本文选题：微紫青霉　切入点：酸性木聚糖酶　出处：《食品科学》2017年14期 　论文类型：期刊论文

【摘要】：基于酸性木聚糖酶在饲料及酿酒行业良好的应用前景,通过基因组步移的方法克隆得到微紫青霉酸性木聚糖酶的全长基因xynA,然后采取重叠延伸PCR技术进行内含子的切除获得xynA的cDNA序列,并对其进行了生物信息学分析。序列分析结果显示xynA基因全长720 bp,内含子63 bp,cDNA全长657 bp。推测该木聚糖酶编码信号肽28个氨基酸,成熟肽190个氨基酸;预测该蛋白为分子质量20.61 kD、等电点7.0的亲水性蛋白,且分子内不含二硫键。与其他真菌来源的GH11族耐酸性木聚糖酶进行序列比对,结果显示该酶的相应位置具有特征天冬氨酸残基Asp,且具有糖苷水解酶11族的保守区域特征以及典型的"右手半握"状结构,重组木聚糖酶基因xynA能够在大肠杆菌中成功表达,比酶活力达220.5 U/mg。
[Abstract]:Based on the good application prospect of acid xylanase in feed and wine industry, The full-length gene XynAof acid xylanase of Penicillium microphylla was cloned by genomic step, and the cDNA sequence of xynA was obtained by excision of intron by overlapping extension PCR. The sequence analysis showed that the full length of xynA gene was 720bpand the full length of intron 63bp cDNA was 657bp. it was deduced that the xylanase encode signal peptide of 28 amino acids and mature peptide of 190 amino acids. The protein was predicted to be a hydrophilic protein with a molecular weight of 20.61 kD and an isoelectric point of 7.0, and no disulfide bond was found in the molecule. The protein was sequenced to GH11 group acidtolerant xylanase from other fungi. The results showed that the enzyme had the characteristic aspartic acid residue AspS, the conserved region of the glycoside hydrolase group 11 and the typical right-hand half-grip structure. The recombinant xylanase gene xynA could be successfully expressed in Escherichia coli. The specific enzyme activity was 220.5Umg.
【作者单位】：北京工商大学北京食品营养与人类健康高精尖创新中心;北京工商大学北京市食品添加剂工程技术研究中心;北京工商大学食品学院;
【基金】：国家自然科学基金面上项目(31371723)
【分类号】：Q78

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