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

Abstract

Peroxisomes are ubiquitous organelles that enclose catalases, fatty acid-oxidizing enzymes, and a variety of proteins involved in different cellular processes. Interestingly, the late enzymes involved in penicillin biosynthesis, and the isopenicillin N epimerization enzymes involved in cephalosporin biosynthesis are located inside peroxisomes in the producer fungi Penicillium chrysogenum and Acremonium chrysogenum. Peroxisome proteins are targeted to those organelles by peroxisomal targeting signals located at the C-terminus (PTS1) or near the N-terminal end (PTS2) of those proteins. Peroxisomal membrane proteins (PMPs) are largely recruited by the interaction with specific sequences in the Pex19 protein. The compartmentalization into peroxisomes of several steps of the biosynthesis of penicillin, cephalosporin, and other secondary metabolites raises the question of how the precursors and/or intermediates of the biosynthesis of β -lactam antibiotics are transported into peroxisomes and the mechanisms of secretion of the final products (penicillin or cephalosporin) from peroxisomes to the extracellular medium. Recent advances in peroxisome proteomics, immunoelectron microscopy, and fluorescence labeling have shown that the transport of these intermediates is mediated by membrane proteins of the major facilitator superfamily class (drug/H + antiporters) containing 12 transmembrane-spanning domains (TMS). In some cases, the transport of the substrates (e.g., fatty acids) or intermediates may be mediated by ATP-binding cassette (ABC) transporters. Knowledge on the transport and secretion mechanisms is of paramount importance to understand the complex mechanisms of cell differentiation and their crosstalk with the biosynthesis of different secondary metabolites that act as biochemical signals between the producer cells and also as communication signals with competing microorganisms (e.g., antimicrobial agents or plant elicitors).

The inducers 1,3-diaminopropane and spermidine cause the reprogramming of metabolism in Penicillium chrysogenum, leading to multiple vesicles and penicillin overproduction

“…This heteromultimeric enzyme couples the energy of ATP hydrolysis to proton transport across intracellular and plasma membranes of eukaryotic cells using the H + gradient to energize the transport of solutes, to maintain a constant intracellular pH, and to provide a signal for intravesicular processes [17,90]. Several carriers, such as those involved in nutrient uptake, utilize the electrochemical gradient of protons (or proton motive force) that exists across the plasma membrane to energize transport.…”

“…Another finding that may be related to this protein is the positive effect that 1,3-DAP exerts on the formation of vesicles. Cargo vesicles formed in the ER have been reported to play an important role in the biosynthesis of aflatoxins [16,17]. Interestingly, one of the enzymes found in these vesicles is spermidine synthase [91].…”

“…Recent evidence in several filamentous fungi revealed an important role of endoplasmic reticulum (ER)-derived vesicles in the production of several secondary metabolites, including aflatoxins in Aspergillus species [15][16][17].…”

Transport systems, intracellular traffic of intermediates and secretion of β-lactam antibiotics in fungi

“…Expression of genes encoding secondary metabolites in gene clusters is frequently coregulate [35,63] what favours the formation of equimolecular amounts of the different biosynthetic enzymes [46] and also of the correct amounts of regulatory proteins encoded by genes that are frequently, although not always, situated in the corresponding gene clusters [36]. In the last decades significant evidence on the temporal organization of expression of different gene in a given gene cluster and also of subcellular localization of the biosynthetic enzymes has been published [52,63,73,74]. Even before the arrival of the omics era several examples of sequential formation of intermediates and final products of a biosynthetic pathway were described.…”

Penicillins

“…It is now well established that peroxisomes are required for effi cient penicillin biosynthesis in P. chrysogenum [ 60 ] and that the penicillin biosynthetic pathway is compartmentalized between the cytosol and peroxisomes [ 24 ]. The distinct subcellular organization of penicillin biosynthesis implies transport of enzymes, precursors, intermediates, and products through these compartments [ 61 ].…”