Microbiological ring expansion of penicillin N

Area of Science:

• Microbiology
• Biochemistry
• Enzymology

Background:

• Cephalosporium acremonium is a fungus known for producing antibiotics.
• Penicillin N and deacetoxycephalosporin C are important beta-lactam antibiotics.
• Understanding the biosynthetic pathways is crucial for optimizing antibiotic production.

Purpose of the Study:

• To isolate and characterize a mutant of Cephalosporium acremonium with enhanced penicillin N production.
• To investigate the enzymatic conversion of penicillin N to deacetoxycephalosporin C.
• To elucidate the properties of the ring-expanding enzyme system involved in cephalosporin biosynthesis.

Main Methods:

• Isolation and characterization of a high-penicillin N producing mutant of Cephalosporium acremonium.
• Enzymatic conversion assays using cell-free extracts from various C. acremonium mutants.
• Purification and characterization of penicillin N.
• Enzyme preparation via ether-treatment, sonication, or osmotic shock.

Main Results:

• A mutant strain overproducing penicillin N was successfully isolated.
• Penicillin N was enzymatically converted to deacetoxycephalosporin C using enzyme systems from C. acremonium.
• The ring-expanding enzyme activity was found in mutants lacking cephem production, including the penicillin N overproducer.
• The enzyme system requires ATP and functions as a 2-oxoglutarate dependent dioxygenase.

Conclusions:

• The ability of a cephem-negative mutant to convert penicillin N to deacetoxycephalosporin C suggests the ring-expanding enzyme complex is functional.
• The overproduction of penicillin N in this mutant is not due to a block in the ring-expanding enzyme.
• The characterization of the enzyme system provides insights into the biosynthesis of cephalosporins.