Related Experiment Video
Updated: Jul 17, 2026
From a Natural Product to Its Biosynthetic Gene Cluster: A Demonstration Using Polyketomycin from Streptomyces diastatochromogenes Tü6028
Published on: January 13, 2017
Cloning and characterization of penicillin V acylase from Streptomyces mobaraensis
Demin Zhang1, Mayuko Koreishi, Hiroyuki Imanaka
1Department of Bioscience and Biotechnology, Faculty of Engineering, Okayama University, 3-1-1, Tsushima-Naka, Okayama 700-8530, Japan.
Abstract:
We report on the molecular cloning and characterization of penicillin V acylase (PVA) from an actinomycete, Streptomyces mobaraensis (Sm-PVA), which was originally isolated as an acylase that efficiently hydrolyzes the amide bond of various N-fatty-acyl-l-amino acids and N-fatty-acyl-peptides as well as capsaicin (8-methyl-N-vanillyl-6-nonenamide). In addition, the purified Sm-PVA hydrolyzed penicillin V with the highest activity (k(cat)) among the PVAs so far reported, penicillin G, and 2-nitro-5-phenoxyacetamide benzoic acid. The BLAST search revealed that the Sm-PVA precursor is composed of a polypeptide that is characteristic of enzymes belonging to the beta-lactam acylase family with four distinct segments; a signal sequence (43 amino acids), an alpha subunit (173 amino acids), a linker peptide (28 amino acids), and a beta subunit (570 amino acids). The mature, active Sm-PVA is a heterodimeric protein with alpha and beta subunits, in contrast to PVAs isolated from Bacillus sphaericus and B. subtilis, which have a homotetrameric structure. The amino acid sequence of Sm-PVA showed identities to PVA from S. lavendulae, N-acylhomoserine lactone-degrading acylase from Streptomyces sp., cyclic lipopeptide acylase from Streptomyces sp., and aculeacin A acylase from Actinoplanes utahensis with 68, 67, 67, and 41% identities, respectively.
Insights
We cloned and characterized penicillin V acylase (PVA) from Streptomyces mobaraensis. This enzyme exhibits high activity, hydrolyzing penicillin V and other substrates, with a unique heterodimeric structure.
Area of Science:
- Enzymology
- Molecular Biology
- Biochemistry
Background:
- Penicillin V acylase (PVA) is crucial for antibiotic modification.
- Actinomycetes are a source of novel enzymes with industrial applications.
Purpose of the Study:
- To molecularly clone and characterize penicillin V acylase (PVA) from Streptomyces mobaraensis (Sm-PVA).
- To investigate the substrate specificity and structural properties of Sm-PVA.
Main Methods:
- Molecular cloning of the Sm-PVA gene.
- Enzyme purification and activity assays.
- Bioinformatic analysis including BLAST search and sequence alignment.
Main Results:
- Sm-PVA demonstrated high catalytic activity (kcat) for penicillin V, surpassing previously reported PVAs.
- The enzyme efficiently hydrolyzed various N-fatty-acyl-l-amino acids, N-fatty-acyl-peptides, capsaicin, penicillin G, and 2-nitro-5-phenoxyacetamide benzoic acid.
- Sm-PVA possesses a heterodimeric structure (alpha and beta subunits), distinct from the homotetrameric PVAs of Bacillus species.
Conclusions:
- Streptomyces mobaraensis is a source of a highly active penicillin V acylase.
- The unique heterodimeric structure of Sm-PVA differentiates it from other known PVAs.
- Sm-PVA has potential applications in pharmaceutical and biotechnological processes.
More Related Videos
05:06Characterizing Multidrug Efflux Systems in Acinetobacter baumannii Using an Efflux-Deficient Bacterial Strain and a Single-Copy Gene Expression System
Published on: January 5, 2024
07:59A High-Yield Streptomyces Transcription-Translation Toolkit for Synthetic Biology and Natural Product Applications
Published on: September 10, 2021
