title: Cloning a cuticle-degrading serine protease gene with biologic control function from Beauveria brongniartii and its expression in Escherichia coli.
abstract: Beauveria brongniartii extracellular subtilisin-like serine protease (Pr1) is one of the most virulent factors by virtue of its activity against insect cuticles. The Pr1 cDNA was cloned using the switching mechanism at the 5' end of the RNA transcript and rapid amplification of cDNA ends. The 1732-bp fragment of genomic DNA containing the predicted open-reading frame of the Pr1 gene was cloned by polymerase chain reaction and sequenced. The Pr1 cDNA is 1550 bp and contains an 1140-bp ORF. The deduced amino-acid sequence of the protein shows identity to that of proteinase K from Tritirachium album (62%), Pr1 from Metarhizium nisopliae (67%), and Pr1 from B. bassiana (76%). The Pr1 protein with an N-terminal fusion to the six-histidine tag was expressed in Escherichia coli as inclusion bodies with the expression vector pBV220. Sodium dodecylsulsulfate-polyacrylamide gel electrophoresis clearly revealed expressed product. The Pr1 protein was purified and refolded and had proteolytic activity of 0.288 U mg(-1).
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, Peoples Republic of China.
authors.forename: Juzheng
authors.initials: J
authors.lastname: Sheng
authors.forename: Kun
authors.initials: K
authors.lastname: An
authors.forename: Chunsheng
authors.initials: C
authors.lastname: Deng
authors.forename: Wenjing
authors.initials: W
authors.lastname: Li
authors.forename: Xiaoming
authors.initials: X
authors.lastname: Bao
authors.forename: Dewen
authors.initials: D
authors.lastname: Qiu
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D005800:Genes, Fungal; D016681:Genome, Fungal; D006999:Hypocreales / Q000201:enzymology* / Q000235:genetics; D007313:Insecta / Q000382:microbiology; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D012697:Serine Endopeptidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D037521:Virulence Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D012697:Serine Endopeptidases