Abstract.
Non-ribosomal peptide synthetases, polyketides and fatty acid synthetases have a modular organisation of multi-enzymatic activities. In all of them, the acyl or peptidyl carrier proteins have 4'-phosphopantetheine (P-pant) as an essential prosthetic group. This is added by 4'-phosphopantetheinyl transferases (PPTases) that derive the P-pant group from coenzyme A. While many PPTases of varying specificity have now been isolated from a number of bacteria, a filamentous fungal PPTase has yet to be characterised. Through database searching of the Aspergillus fumigatus genome sequence against Sfp from Bacillus subtilis, we identified a unique sequence which appears to encode a PPTase, as deduced from conserved residues considered important in PPTases. The PPTase candidate was used to search the NCBI data base and an unexpected homologue in A. nidulans was identified as npgA. Mutations in this gene (cfwA/npgA) were identified previously as leading to defects in growth and pigmentation. To test whether the temperature-sensitive cfwA2 mutation impairs penicillin biosynthesis, which is dependent on the δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine synthetase, bioassays with B. calidolactis were set up at permissive and non-permissive temperatures. The cfwA2 mutant did not produce penicillin at the non-permissive temperature. Since no other PPTase homologue has been detected in the A. fumigatus genome to date, the data suggest that a single enzyme may be able to transfer the cofactor to a broad range of enzymes with acyl or peptidyl carrier protein domains.
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References
Aguirre J, Ortiz R, Clutterbuck J, Tapia R, Cardenas M (1993) vegA and cfwA define two new developmental genes in Aspergillus nidulans. Fungal Genet Newsl 40a:68
Aramayo R, Timberlake WE (1990) Sequence and molecular structure of the Aspergillus nidulans yA (laccase I) gene. Nucleic Acids Res 18:3415
Ballance DJ, Turner G (1985) Development of a high-frequency transforming vector for Aspergillus nidulans. Gene 36:321–331
Brakhage AA, Browne P, Turner G (1992) Regulation of Aspergillus nidulans penicillin biosynthesis and penicillin biosynthesis genes acvA and ipnA by glucose. J Bacteriol 174:3789–3799
Cosmina P, Rodriguez F, Ferra F de, Grandi G, Perego M, Venema G. Sinderen D van (1993) Sequence and analysis of the genetic locus responsible for surfactin synthesis in Bacillus subtilis. Mol Microbiol 8:821–831
Ehmann DE, Gehring AM, Walsh CT (1999) Lysine biosynthesis in Saccharomyces cerevisiae: mechanism of alpha-aminoadipate reductase (Lys2) involves posttranslational phosphopantetheinylation by Lys5. Biochemistry 38:6171–6177
Fichtlscherer F, Wellein C, Mittag M, Schweizer E (2000) A novel function of yeast fatty acid synthetase. Subunit alpha is capable of self-pantetheinylation. Eur J Biochem 267:2666–2671
Han YJ, Han DM (1993) Isolation and characterization of null pigment mutant in Aspergillus nidulans. Korean J Genet 15:1–10
Kennedy J, Turner G (1996) δ–(l-α-Aminoadipyl)-l-cysteinyl-d-valine synthetase is a rate limiting enzyme for penicillin production in Aspergillus nidulans. Mol Gen Genet 253:189–197
Kim J-M, Han D-M, Chae K-S, Chae K-S, Jahng K-Y (2001) Isolation and characterization of the npgA gene involved in pigment formation in Aspergillus nidulans. Fungal Genet Newsl 48:52
Kleinkauf H, Döhren H von (1996) A nonribosomal system of peptide biosynthesis. Eur J Biochem 236:335–351
Lambalot RH, Gehring AM, Flugel RS, Zuber P, LaCelle M, Marahiel MA, Reid R, Khosla C, Walsh CT (1996) A new enzyme superfamily—the phosphopantetheinyl transferases. Chem Biol 3:923–936
MacCabe AP, Liempt H van, Palissa H, Unkles SE, Riach MBR, Pfeifer E, Döhren H von, Kinghorn JR (1991) δ–(l-α-Aminoadipyl)-l-cysteinyl-d-valine synthetase from Aspergillus nidulans—molecular characterisation of the acvA gene encoding the first enzyme of the penicillin biosynthetic pathway. J Biol Chem 266:12646–12654
Mootz HD, Schörgendorfer K, Marahiel MA (2002) Functional characterisation of 4'-phosphopantetheinyl transferase genes of bacterial and fungal origin by complementation of Saccharomyces cerevisiae lys5. FEMS Microbiol Lett 213:51–57
Praphanphoj V, Sacksteder KA, Gould SJ, Thomas GH, Geraghty MT (2001) Identification of the α-aminoadipic semialdehyde dehydrogenase-phosphopantetheinyl transferase gene, the human ortholog of the yeast LYS5 gene. Mol Genet Metab 72:336–342
Reuter K, Mofid MR, Marahiel MA, Ficner R (1999) Crystal structure of the surfactin synthetase-activating enyme Sfp: a prototype of the 4'-phosphopantetheinyl transferase superfamily. EMBO J 18:6823–6831
Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
Schneider R, Massow M, Lisowsky T, Weiss H (1995) Different respiratory-defective phenotypes of Neurospora crassa and Saccharomyces cerevisiae after inactivation of the gene encoding the mitochondrial acyl carrier protein. Curr Genet 29:10–17
Smith DJ, Bull JH, Edwards J, Turner G (1989) Amplification of the isopenicillin N synthetase gene in a strain of Penicillium chrysogenum producing high levels of penicillin. Mol Gen Genet 216:492–497
Smith DJ, Earl AJ, Turner G (1990) The multifunctional peptide synthetase performing the first step of penicillin biosynthesis is a 421073 dalton protein similar to Bacillus brevis peptide antibiotic synthetases. EMBO J 9:2743–2750
Stuible HP, Meier S, Wagner C, Hannappel E, Schweizer E (1998) A novel phosphopantetheine:protein transferase activating yeast mitochondrial acyl carrier protein. J Biol Chem 273:22334–22339
Watanabe A, Fujii I, Sankawa U, Mayorga ME, Timberlake WE, Ebizuka Y (1999) Re-identification of Aspergillus nidulans wA gene to code for a polyketide synthetase of naphthopyrone. Tetrahedron Lett 40:91–94
Weber G, Schorgendorfer K, Schneider-Scherzer E, Leitner E (1994) The peptide synthetase catalyzing cyclosporine production in Tolypocladium niveum is encoded by a giant 45.8-kilobase open reading frame. Curr Genet 26:120–125
Acknowledgements.
We thank Dr. J. Clutterbuck for providing AJC12:36. This work was supported in part (D.K.P.) by EU grant QTRK3-1999-00729 (Eurofung).
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Keszenman-Pereyra, D., Lawrence, S., Twfieg, ME. et al. The npgA/cfwA gene encodes a putative 4'-phosphopantetheinyl transferase which is essential for penicillin biosynthesis in Aspergillus nidulans . Curr Genet 43, 186–190 (2003). https://doi.org/10.1007/s00294-003-0382-7
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DOI: https://doi.org/10.1007/s00294-003-0382-7