scholarly journals Omics Approaches Applied to Penicillium chrysogenum and Penicillin Production: Revealing the Secrets of Improved Productivity

Genes ◽  
2020 ◽  
Vol 11 (6) ◽  
pp. 712
Author(s):  
Carlos García-Estrada ◽  
Juan F. Martín ◽  
Laura Cueto ◽  
Carlos Barreiro
Keyword(s):  
Synthetic Biology ◽  
Secondary Metabolites ◽  
Penicillium Chrysogenum ◽  
Molecular Mechanisms ◽  
Penicillin Biosynthesis ◽  
Biological Processes ◽  
Penicillin Production ◽  
Bioactive Secondary Metabolites ◽  
Industrial Strains ◽  
Points Of View

Penicillin biosynthesis by Penicillium chrysogenum is one of the best-characterized biological processes from the genetic, molecular, biochemical, and subcellular points of view. Several omics studies have been carried out in this filamentous fungus during the last decade, which have contributed to gathering a deep knowledge about the molecular mechanisms underlying improved productivity in industrial strains. The information provided by these studies is extremely useful for enhancing the production of penicillin or other bioactive secondary metabolites by means of Biotechnology or Synthetic Biology.

scholarly journals Characterization of an Autoinducer of Penicillin Biosynthesis in Penicillium chrysogenum

2011 ◽  
Vol 77 (16) ◽  
pp. 5688-5696 ◽  
Author(s):  
Jorge Martín ◽  
Carlos García-Estrada ◽  
Ángel Rumbero ◽  
Eliseo Recio ◽  
Silvia M. Albillos ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Penicillium Chrysogenum ◽  
Molecular Mechanisms ◽  
Biological Activities ◽  
Northern Analysis ◽  
Penicillin Biosynthesis ◽  
Control Of Gene Expression ◽  
Content Type ◽  
Direct Testing

ABSTRACTFilamentous fungi produce an impressive variety of secondary metabolites; many of them have important biological activities. The biosynthesis of these secondary metabolites is frequently induced by plant-derived external elicitors and appears to also be regulated by internal inducers, which may work in a way similar to that of bacterial autoinducers. The biosynthesis of penicillin inPenicillium chrysogenumis an excellent model for studying the molecular mechanisms of control of gene expression due to a good knowledge of the biochemistry and molecular genetics of β-lactam antibiotics and to the availability of its genome sequence and proteome. In this work, we first developed a plate bioassay that allows direct testing of inducers of penicillin biosynthesis using single colonies ofP. chrysogenum. Using this bioassay, we have found an inducer substance in the conditioned culture broths ofP. chrysogenumandAcremonium chrysogenum. No inducing effect was exerted by γ-butyrolactones, jasmonic acid, or the penicillin precursor δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine. The conditioned broth induced penicillin biosynthesis and transcription of thepcbAB,pcbC, andpenDEgenes when added at inoculation time, but its effect was smaller if added at 12 h and it had no effect when added at 24 h, as shown by Northern analysis andlacZreporter studies. The inducer molecule was purified and identified by mass spectrometry (MS) and nuclear magnetic resonance (NMR) as 1,3-diaminopropane. Addition of pure 1,3-diaminopropane stimulated the production of penicillin by about 100% compared to results for the control cultures. Genes for the biosynthesis of 1,3-diaminopropane have been identified in theP. chrysogenumgenome.

scholarly journals Increased Penicillin Production in Penicillium chrysogenum Production Strains via Balanced Overexpression of Isopenicillin N Acyltransferase

2012 ◽  
Vol 78 (19) ◽  
pp. 7107-7113 ◽  
Author(s):  
Stefan S. Weber ◽  
Fabiola Polli ◽  
Rémon Boer ◽  
Roel A. L. Bovenberg ◽  
Arnold J. M. Driessen
Keyword(s):  
Penicillium Chrysogenum ◽  
Phenylacetic Acid ◽  
Strain Improvement ◽  
Single Copy ◽  
Limiting Factor ◽  
Penicillin Biosynthesis ◽  
Penicillin Production ◽  
Content Type ◽  
Coa Ligase ◽  
Isopenicillin N

ABSTRACTIntense classical strain improvement has yielded industrialPenicillium chrysogenumstrains that produce high titers of penicillin. These strains contain multiple copies of the penicillin biosynthesis cluster encoding the three key enzymes: δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine synthetase (ACVS), isopenicillin N synthase (IPNS), and isopenicillin N acyltransferase (IAT). The phenylacetic acid coenzyme A (CoA) ligase (PCL) gene encoding the enzyme responsible for the activation of the side chain precursor phenylacetic acid is localized elsewhere in the genome in a single copy. Since the protein level of IAT already saturates at low cluster copy numbers, IAT might catalyze a limiting step in high-yielding strains. Here, we show that penicillin production in high-yielding strains can be further improved by the overexpression of IAT while at very high levels of IAT the precursor 6-aminopenicillic acid (6-APA) accumulates. Overproduction of PCL only marginally stimulates penicillin production. These data demonstrate that in high-yielding strains IAT is the limiting factor and that this limitation can be alleviated by a balanced overproduction of this enzyme.

scholarly journals Nonlinear Biosynthetic Gene Cluster Dose Effect on Penicillin Production by Penicillium chrysogenum

2010 ◽  
Vol 76 (21) ◽  
pp. 7109-7115 ◽  
Author(s):  
Jeroen G. Nijland ◽  
Bjorg Ebbendorf ◽  
Marta Woszczynska ◽  
Rémon Boer ◽  
Roel A. L. Bovenberg ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Penicillium Chrysogenum ◽  
Copy Number ◽  
Biosynthetic Gene Cluster ◽  
Dose Effect ◽  
Penicillin Biosynthesis ◽  
Biosynthetic Gene ◽  
Penicillin Production ◽  
Copy Numbers ◽  
Isopenicillin N

ABSTRACT Industrial penicillin production levels by the filamentous fungus Penicillium chrysogenum increased dramatically by classical strain improvement. High-yielding strains contain multiple copies of the penicillin biosynthetic gene cluster that encodes three key enzymes of the β-lactam biosynthetic pathway. We have analyzed the gene cluster dose effect on penicillin production using the high-yielding P. chrysogenum strain DS17690 that was cured from its native clusters. The amount of penicillin V produced increased with the penicillin biosynthetic gene cluster number but was saturated at high copy numbers. Likewise, transcript levels of the biosynthetic genes pcbAB [δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine synthetase], pcbC (isopenicillin N synthase), and penDE (acyltransferase) correlated with the cluster copy number. Remarkably, the protein level of acyltransferase, which localizes to peroxisomes, was saturated already at low cluster copy numbers. At higher copy numbers, intracellular levels of isopenicillin N increased, suggesting that the acyltransferase reaction presents a limiting step at a high gene dose. Since the number and appearance of the peroxisomes did not change significantly with the gene cluster copy number, we conclude that the acyltransferase activity is limiting for penicillin biosynthesis at high biosynthetic gene cluster copy numbers. These results suggest that at a high penicillin production level, productivity is limited by the peroxisomal acyltransferase import activity and/or the availability of coenzyme A (CoA)-activated side chains.

α-Aminoadipate pool concentration and penicillin biosynthesis in strains of Penicillium chrysogenum

1986 ◽  
Vol 32 (6) ◽  
pp. 473-480 ◽  
Author(s):  
W. M. Jaklitsch ◽  
W. Hampel ◽  
M. Röhr ◽  
C. P. Kubicek ◽  
G. Gamerith
Keyword(s):  
Amino Acid ◽  
Penicillium Chrysogenum ◽  
Intracellular Concentration ◽  
Lysine Biosynthesis ◽  
Penicillin Biosynthesis ◽  
Penicillin Production ◽  
Saccharopine Dehydrogenase ◽  
Two Factors ◽  
Homocitrate Synthase ◽  
Rate Limiting

Intracellular amino acid pools in four Penicillium chrysogenum strains, which differed in their ability to produce penicillin, were determined under conditions supporting growth without penicillin production and under conditions supporting penicillin production. A significant correlation between the rate of pencillin production and the intracellular concentration of α-aminoadipate was observed, which was not shown with any other amino acid in the pool. In replacement cultivation, penicillin production was stimulated by α-aminoadipate, but not by valine or cysteine. Exogenously added α-aminoadipate (2 or 3 mM) maximally stimulated penicillin synthesis in two strains of different productivity. Under these conditions intracellular concentrations of α-aminoadipate were comparable in the two strains in spite of the higher rate of penicillin production in the more productive strain. Results suggest that the lower penicillin titre of strain Q 176 is due to at least two factors: (i) the intracellular concentration of α-aminoadipate is insufficient to allow saturation of any enzyme which is rate limiting in the conversion of α-aminoadipate to penicillin and (ii) the level of an enzyme, which is rate limiting in the conversion of α-aminoadipate to penicillin, is lower in Q 176 (relative to strain D6/1014/A). Results suggest that the intracellular concentration of α-aminoadipate in strain D6/1014/A is sufficiently high to allow saturation of the rate-limiting penicillin biosynthetic enzyme in that strain. The basis of further correlation of intracellular α-aminoadipate concentration and penicillin titre among strains D6/1014/A, P2, and 389/3, the three highest penicillin producers studied here, remains to be established. Preliminary studies which attempted to explain the differences in intracellular α-aminoadipate concentrations in strains Q 176, D6/1014/A, and P2 in terms of differences in activities or kinetics of two enzymes of lysine biosynthesis (homocitrate synthase and saccharopine dehydrogenase) did not reveal differences in those enzymes among the three strains.

Regulation of α-aminoadipate reductase from Penicillium chrysogenum in relation to the flux from α-aminoadipate into penicillin biosynthesis

1992 ◽  
Vol 38 (8) ◽  
pp. 758-763 ◽  
Author(s):  
Ying Lu ◽  
Robert L. Mach ◽  
Karin Affenzeller ◽  
Christian P. Kubicek
Keyword(s):  
Key Words ◽  
Penicillium Chrysogenum ◽  
Reductase Activity ◽  
Lysine Biosynthesis ◽  
Penicillin Biosynthesis ◽  
Penicillin Production ◽  
Producer Strain ◽  
Ph Dependent ◽  
Producer Strains

The activity and regulation of α-aminoadipate reductase in three Penicillium chrysogenum strains (Q176, D6/1014/A, and P2), producing different amounts of penicillin, were studied. The enzyme exhibited decreasing affinity for α-aminoadipate with increasing capacity of the respective strain to produce penicillin. The enzyme from all three strains was inhibited by L-lysine, and the enzyme from the lowest producer, Q176, was least sensitive. Between pH 7.5 and 6.5, inhibition of α-aminoadipate reductase by L-lysine was pH dependent, being more pronounced at lower pH. The highest producer strain, P2, displayed the lowest α-aminoadipate reductase activity at pH 7.0. In Q176, the addition of 0.5–1 mM of exogenous lysine stimulated penicillin formation, whereas the same concentration was ineffective or inhibitory with strains D6/1014/A and P2. The addition of higher (up to 5 mM) lysine concentrations inhibited penicillin production in all three strains. In mutants of P. chrysogenum D6/1014/A, selected for resistance to 20 mM α-aminoadipate, highest penicillin production was observed in those strains whose α-aminoadipate reductase was most strongly inhibited by L-lysine. The results support the conclusion that the in vivo activity of α-aminoadipate reductase from superior penicillin producer strains of P. chrysogenum is more strongly inhibited by lysine, and that this is related to their ability to accumulate increased amounts of α-aminoadipate, and hence penicillin. Key words: α-aminoadipate, α-aminoadipate reductase, regulation of lysine biosynthesis, penicillin biosynthesis, Penicillium chrysogenum.

scholarly journals Post-translational enzyme modification by the phosphopantetheinyl transferase is required for lysine and penicillin biosynthesis but not for roquefortine or fatty acid formation in Penicillium chrysogenum

2008 ◽  
Vol 415 (2) ◽  
pp. 317-324 ◽  
Author(s):  
Carlos García-Estrada ◽  
Ricardo V. Ullán ◽  
Tania Velasco-Conde ◽  
Ramiro P. Godio ◽  
Fernando Teijeira ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Penicillium Chrysogenum ◽  
Fatty Acid Synthase ◽  
Single Copy ◽  
Lysine Biosynthesis ◽  
Penicillin Biosynthesis ◽  
Penicillin Production ◽  
Phosphopantetheinyl Transferase ◽  
Wide Range ◽  
Multiple Copies

NRPSs (non-ribosomal peptide synthetases) and PKSs (polyketide synthases) require post-translational phosphopantetheinylation to become active. This reaction is catalysed by a PPTase (4′-phosphopantetheinyl transferase). The ppt gene of Penicillium chrysogenum, encoding a protein that shares 50% similarity with the stand-alone large PPTases, has been cloned. This gene is present as a single copy in the genome of the wild-type and high-penicillin-producing strains (containing multiple copies of the penicillin gene cluster). Amplification of the ppt gene produced increases in isopenicillin N and benzylpenicillin biosynthesis. A PPTase-defective mutant (Wis54-PPT−) was obtained. It required lysine and lacked pigment and penicillin production, but it still synthesized normal levels of roquefortine. The biosynthesis of roquefortine does not appear to involve PPTase-mediated modification of the synthesizing enzymes. The PPT− mutant did not require fatty acids, which indicates that activation of the fatty acid synthase is performed by a different PPTase. Complementation of Wis54-PPT− with the ppt gene restored lysine biosynthesis, pigmentation and penicillin production, which demonstrates the wide range of processes controlled by this gene.

Bioactive Secondary Metabolites from Geosmithia langdonii

Planta Medica ◽  
2013 ◽  
Vol 79 (10) ◽  
Author(s):  
LG Malak ◽  
DW Bishay ◽  
AM Abdel-baky ◽  
AM Moharram ◽  
SJ Cutler ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Bioactive Secondary Metabolites

Screening bioactive secondary metabolites from Costa Rican enviromental microbial communities

Planta Medica ◽  
2015 ◽  
Vol 81 (11) ◽  
Author(s):  
JJ Araya ◽  
M Chavarría ◽  
A Pinto-Tomás ◽  
C Murillo ◽  
L Uribe ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Microbial Communities ◽  
Costa Rican ◽  
Bioactive Secondary Metabolites
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