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Overexpression of Penicillin V Acylase from Streptomyces lavendulae and Elucidation of Its Catalytic Residues

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Torres Bacete, Jesús and Hormigo, Daniel and Torres Guzmán, Raquel and Arroyo Sánchez, Miguel and Castillón, María Pilar and García, José Luis and Acebal Sarabia, Carmen and Mata Riesco, Isabel de la (2015) Overexpression of Penicillin V Acylase from Streptomyces lavendulae and Elucidation of Its Catalytic Residues. Applied and Environmental Microbiology, 81 (4). pp. 1225-1233. ISSN 0099-2240

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Abstract

The pva gene from Streptomyces lavendulae ATCC 13664, encoding a novel penicillin V acylase (SlPVA), has been isolated and characterized. The gene encodes an inactive precursor protein containing a secretion signal peptide that is activated by two internal autoproteolytic cleavages that release a 25-amino-acid linker peptide and two large domains of 18.79 kDa (_-subunit) and 60.09 kDa (_-subunit). Based on sequence alignments and the three-dimensional model of SlPVA, the enzyme contains a hydrophobic pocket involved in catalytic activity, including Ser_1, His_23, Val_70, and Asn_272, which were confirmed by site-directed mutagenesis studies. The heterologous expression of pva in S. lividans led to the production of an extracellularly homogeneous heterodimeric enzyme at a 5-fold higher concentration (959 IU/liter) than in the original host and in a considerably shorter time. According to the catalytic properties of SlPVA, the enzyme must be classified as a new member of the Ntn-hydrolase superfamily, which belongs to a novel subfamily of acylases that recognize substrates with long hydrophobic acyl chains and have biotechnological applications in semisynthetic antifungal production.


Item Type:Article
Uncontrolled Keywords:Microbiology; Penicillin V Acylase; Streptomyces lavendulae
Subjects:Medical sciences > Biology
Medical sciences > Biology > Molecular biology
Medical sciences > Biology > Microbiology
ID Code:43288
Deposited On:13 Jun 2017 16:02
Last Modified:14 Jun 2017 07:38

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