Regulation and compartmentalization of β‐lactam biosynthesis

Abstract

SummaryPenicillins and cephalosporins are β‐lactam antibiotics widely used in human medicine. The biosynthesis of these compounds starts by the condensation of the amino acids l‐α‐aminoadipic acid, l‐cysteine and l‐valine to form the tripeptide δ‐l‐α‐aminoadipyl‐l‐cysteinyl‐d‐valine catalysed by the non‐ribosomal peptide ‘ACV synthetase’. Subsequently, this tripeptide is cyclized to isopenicillin N that in Penicillium is converted to hydrophobic penicillins, e.g. benzylpenicillin. In Acremonium and in streptomycetes, isopenicillin N is later isomerized to penicillin N and finally converted to cephalosporin. Expression of genes of the penicillin (pcbAB, pcbC, pendDE) and cephalosporin clusters (pcbAB, pcbC, cefD1, cefD2, cefEF, cefG) is controlled by pleitropic regulators including LaeA, a methylase involved in heterochromatin rearrangement. The enzymes catalysing the last two steps of penicillin biosynthesis (phenylacetyl‐CoA ligase and isopenicillin N acyltransferase) are located in microbodies, as shown by immunoelectron microscopy and microbodies proteome analyses. Similarly, the Acremonium two‐component CefD1–CefD2 epimerization system is also located in microbodies. This compartmentalization implies intracellular transport of isopenicillin N (in the penicillin pathway) or isopenicillin N and penicillin N in the cephalosporin route. Two transporters of the MFS family cefT and cefM are involved in transport of intermediates and/or secretion of cephalosporins. However, there is no known transporter of benzylpenicillin despite its large production in industrial strains.

Transport of substrates into peroxisomes: the paradigm of β-lactam biosynthetic intermediates

“…Authentic benzylpenicillin transporters in the peroxisomal or in the fungal membrane have not been scientifically confirmed. Alternatively, the fusion of peroxisomes with secretory vesicles that may unload the penicillin formed by an exocytosis mechanism has been proposed (94,116,138) . This vesicle-mediated mechanism may be different from the known pexophagy because a P. chrysogenum mutant defective in pexophagy produces normal or even higher levels of penicillin due to the preservation of peroxisomes and the biosynthetic enzymes (4) .…”

“…The secretion of penicillin from the peroxisome to the medium is a matter of debate (5,9,116,145) . Authentic benzylpenicillin transporters in the peroxisomal or in the fungal membrane have not been scientifically confirmed.…”

“…In the A. chrysogenum ATCC 48272 strain, the pcbAB , pcbC , cefD1 , and cefD2 biosynthetic genes and the secretion/translocation genes cefT , cefM , and cefP are located on chromosome VII in the so-called early cluster (89, 127 -130) . This ' early ' cluster contains all the genetic information for the biosynthesis of the IPN and PenN antibiotics (the latter is efficiently secreted), and therefore, it can be considered as a complete gene cluster for these penicillins (5,116) . The late cephalosporin biosynthetic genes cefEF and cefG are located in a second cluster, named ' late cluster ' , on chromosome I (127) .…”

“…Cephalosporin C belongs to the class of broad-spectrum antibiotics obtained from cultures of the filamentous fungus A. chrysogenum (114,115) . The cephalosporin C biosynthetic pathway begins with the non-ribosomal condensation of the three precursor amino acids l -α -aminoadipic acid, l -cysteine, and l -valine to form the tripeptide ACV (116) . This reaction is carried out by the ACV synthetase (117,118) , which is encoded by the pcb AB gene (119) .…”

Proteomics Methodology Applied to the Analysis of Filamentous Fungi - New Trends for an Impressive Diverse Group of Organisms

“…An interesting phenomenon that has been observed in the P. chrysogenum secretome is the fact that some extracellular proteins were also previously identified in the microbody matrix of this fungus (Kiel et al, 2009). Therefore, an alternative explanation given to the presence of intracellular proteins in the culture broth was the selective autophagic degradation of peroxisomes (pexophagy), which can give a new sense to the strange "moonlighting proteins" as it was explained in P. chrysogenum (Jami et al, 2010b;Martín et al, 2010). It is well known that peroxisome abundance can be rapidly decreased through autophagic pathways, which selectively degrade peroxisomes by fusion to lysosomes or vacuoles (Oku and Sakai, 2010).…”