title: Cloning, recombinant expression and characterization of a new phytase from Penicillium chrysogenum.
abstract: The phy gene, which encodes a phytase in Penicillium chrysogenum CCT 1273, was cloned into the vector pAN-52-1-phy and the resulting plasmid was used for the cotransformation of Penicillium griseoroseum PG63 protoplasts. Among the 91 transformants obtained, 23 were cotransformants. From there, the phytase activity of these 23 transformants was evaluated and P. griseoroseum T73 showed the highest. The recombinant strain P. griseoroseum T73 contained the phy gene integrated in at least three sites of the genome and showed a 5.1-fold increase in phytase activity in comparison to the host strain (from 0.56 ± 0.2 to 2.86 ± 0.4 U μg protein(-1)). The deduced PHY protein has 483 amino acids; an isoelectric point (pI) higher than that reported for phytases from filamentous fungi (7.6); higher activity at pH 2.0 (73%), pH 5.0 (100%) and 50 °C; and is stable at pH values 3.0-8.0 and temperatures 70-80 °C. PHY produced by the recombinant strain P. griseoroseum T73 was stable after four weeks of storage at -20, 8 and 25 °C and was effective in releasing Pi, especially from soybeans. The data presented here show that P. griseoroseum is a successful host for expression of heterologous protein and suggest the potential use of PHY in the animal nutrition industry.
authors.affiliation: Department of Microbiology/BIOAGRO, Federal University of Viçosa, Viçosa, MG, Brazil.
authors.forename: Thamy Lívia
authors.initials: TL
authors.lastname: Ribeiro Corrêa
authors.affiliation: Department of Microbiology/BIOAGRO, Federal University of Viçosa, Viçosa, MG, Brazil.
authors.forename: Marisa Vieira
authors.initials: MV
authors.lastname: de Queiroz
authors.affiliation: Department of Microbiology/BIOAGRO, Federal University of Viçosa, Viçosa, MG, Brazil. Electronic address: ezfa@ufv.br.
authors.forename: Elza Fernandes
authors.initials: EF
authors.lastname: de Araújo
mesh_terms: D010832:6-Phytase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004789:Enzyme Activation; D018628:Gene Dosage; D015870:Gene Expression; D008969:Molecular Sequence Data; D010408:Penicillium chrysogenum / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins; D016415:Sequence Alignment; D013816:Thermodynamics
chemical_list: D011994:Recombinant Proteins; D010832:6-Phytase
keywords: Characterization; Molecular cloning; Penicillium chrysogenum; Penicillium griseoroseum PG63; Phytase