一色齿毛菌锰过氧化物酶（MnP）活性检测与MnP1基因的克隆

doi:10.11707/j.1001-7488.20140315

Abstract

Abstract:

Manganese peroxidase (MnP) widely exists in white-rot fungi and it is the main enzyme to degrade lignin. The ligninolytic peroxidase activity of Cerena unicolor stain CB1 collected from Changbai Mountain was firstly detected, result showed that C. unicolor could produce MnP, but no lignin peroxidase (LiP) excreted, and Mn²⁺was not the essential factor for C. unicolor to produce MnP. The highest MnP activity was gained when sawdust was added to LNAS culture solution with Mn²⁺ substrate which was 18.4 U·L^-1. On the basis of MnP activity determination, a MnP full length DNA gene termed Cu-mnp1 (GenBank accession No. JQ782578.1) was cloned in order to study the function of MnP to degrade lignin furtherly. Forward and reverse primers for PCR and nested gene-specific primers for genome walking PCR were designed based on the conserved sequences of the known MnP genes and amplified DNA fragment, all the PCR reactions were conducted by the genomic DNA of the C. unicolor stain CB1 as the template. The full length DNA gene Cu-mnp1 was 4 268 bp containing 16 exons and 15 introns, its promoter region contained diverse cis-acting elements such as TATA-Box, CAAT-Box, SP1, AP1, HSE, and XRE. Its cDNA gene contained 1 092 bp coding region encoding 363 amino acids. BLAST analysis indicated that Cu-mnp1 in the cDNA level showed the highest similarity with mnp2 gene (GenBank accession No. JQ248599.1)from Hericium erinaceum strain CB1 which was 70%. Protein structure analysis showed that Cu-Mnp1 contained 4 disulfide bonds belonging to short MnP.

Key words: Cerena unicolor, ligninolytic enzyme, manganese peroxidase (MnP), gene cloning, genome walking

CLC Number:

S718.81

Yu Cun, Chi Yujie. Detection on Manganese Peroxidase Activity and Cloning of Cu-mnp1 of Cerena unicolor[J]. Scientia Silvae Sinicae, 2014, 50(3): 109-116.

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Detection on Manganese Peroxidase Activity and Cloning of Cu-mnp1 of Cerena unicolor

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