The beta-lactam antibiotic penicillin represents one of the predominantly used antibiotics for the therapy of infectious diseases. It is produced as end product by certain filamentous fungi – most notably Penicillium chrysogenum. Although discovered in 1928, the biosynthesis pathways leading to penicillin have been elucidated much later.

Researchers from the Massachusetts Institute of Technology continued to work on the problem of the chemical and biosynthesis of penicillin, despite the fact that the structure has been completely worked out and mass production of the drug by fermentation processes was quick. In 1957 they finally achieved the total synthesis of natural penicillin.

Three steps of penicillin biosynthesis

Penicillin biosynthesis is often divided into three important steps. The first catalytic step is mediated by delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase (also known as ACV synthetase), which is a high molecular mass enzyme that condenses three leading amino acids, L-alpha-aminoadipate, L-cysteine and L-valine, into the tripeptide ACV.

In the second step, oxidative ring closure of the linear tripeptide results in a formation of a bicyclic ring, i.e. the four-membered beta-lactam ring fused to the five-membered thiazolidine ring that is characteristic for all penicillins. The enzyme employed in this step is isopenicillin N synthase, and the resulting compound is isopenicillin N (IPN) with a weak antibiotic activity, making it the first bioactive intermediate of penicillin biosynthesis pathway.

At this point, the pathway can diverge for the different microorganisms. In the case ofPenicillium (and the other penicillin-producing fungi), the third step represents the exchange of the lateral chain of L-alpha-aminoadipate by a hydrophobic lateral chain; on the other hand, in the genus Acremonium isopenicillin N is converted into penicillin N via a two-enzyme system (acyl-CoA-synthetase and acyl-CoA-racemase).

Genetics of biosynthesis

Genes for beta-lactam antibiotic biosynthesis are found clustered both in filamentous fungi and bacteria. Their organization is common to all the beta-lactam producers, which includes the genes pcbAB for ACV synthetase and pcbC for ACV cyclase. Such linkage of antibiotic-biosynthesis genes is a well-described phenomenon in a myriad of antibiotic-producing organisms.

Expression of genes of the penicillin cluster is controlled by pleitropic regulators including LaeA - a methylase involved in heterochromatin rearrangement. Furthermore, repressors like AnBH1 and VeA are also involved in the regulation.

Key enzymatic steps in penicillin biosynthesis are confined to only two compartments – the cytosol and microbody. Using subcellular fractionation and immuno-electron microscopy, ACV synthetase was localized in the cytosol (together with isopenicillin N synthetase), whereas some of the enzymes pivotal for the third step of penicillin biosynthesis are localized in microbodies.

Penicillin biosynthesis genes are controlled by a complex regulatory network, influenced by factors such as ambient pH, carbon source, amino acids, nitrogen, etc. All aforementioned investigations have contributed to the elucidation of signals leading to the production of penicillin, and thus can be expected to have significant impact on rational strain improvement programs.