Identification and characterization of the biosynthetic gene cluster of polyoxypeptin A, a potent apoptosis inducer

BMC Microbiology

Shuangjun Lin¹  Zixin Deng¹  Meifeng Tao¹  Tingting Huang²  Yemin Wang¹  Yanhua Du¹ 

[1] State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 800 Dongchuan Road, Shanghai 200240, China;Current address: Department of Chemistry at the Scripps Research Institute, Jupiter, FL 33458, USA

关键词: Apoptosis-inducing activity;    Nonribosomal peptide;    Polyketide;    Polyoxypeptin A;    Biosynthesis;   

Others  :  1141995
DOI  :  10.1186/1471-2180-14-30

 received in 2013-11-07, accepted in 2014-02-04,  发布年份 2014

【 摘 要 】

Background

Polyoxypeptin A was isolated from a culture broth of Streptomyces sp. MK498-98 F14, which has a potent apoptosis-inducing activity towards human pancreatic carcinoma AsPC-1 cells. Structurally, polyoxypeptin A is composed of a C₁₅ acyl side chain and a nineteen-membered cyclodepsipeptide core that consists of six unusual nonproteinogenic amino acid residues (N-hydroxyvaline, 3-hydroxy-3-methylproline, 5-hydroxypiperazic acid, N-hydroxyalanine, piperazic acid, and 3-hydroxyleucine) at high oxidation states.

Results

A gene cluster containing 37 open reading frames (ORFs) has been sequenced and analyzed for the biosynthesis of polyoxypeptin A. We constructed 12 specific gene inactivation mutants, most of which abolished the production of polyoxypeptin A and only ΔplyM mutant accumulated a dehydroxylated analogue polyoxypeptin B. Based on bioinformatics analysis and genetic data, we proposed the biosynthetic pathway of polyoxypeptin A and biosynthetic models of six unusual amino acid building blocks and a PKS extender unit.

Conclusions

The identified gene cluster and proposed pathway for the biosynthesis of polyoxypeptin A will pave a way to understand the biosynthetic mechanism of the azinothricin family natural products and provide opportunities to apply combinatorial biosynthesis strategy to create more useful compounds.

【 授权许可】

   
2014 Du et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Umezawa KNK, Uemura T, et al.: Polyoxypeptin isolated from Streptomyces: a bioactive cyclic depsipeptide containing the novel amino acid 3-hydroxy-3-methylproline. Tetrahedron Lett 1998, 39(11):1389-1392.
• [2]Umezawa K, Nakazawa K, Ikeda Y, Naganawa H, Kondo S: Polyoxypeptins A and B produced by Streptomyces: apoptosis-inducing cyclic depsipeptides containing the novel amino acid (2S,3R)-3-hydroxy-3-methylproline. J Org Chem 1999, 64(9):3034-3038.
• [3]Smitka TA, Deeter JB, Hunt AH, Mertz FP, Ellis RM, Boeck LD, Yao RC: A83586C, a new depsipeptide antibiotic. J Antibiot (Tokyo) 1988, 41(6):726-733.
• [4]Grafe U, Schlegel R, Ritzau M, Ihn W, Dornberger K, Stengel C, Fleck WF, Gutsche W, Hartl A, Paulus EF: Aurantimycins, new depsipeptide antibiotics from Streptomyces aurantiacus IMET 43917. Production, isolation, structure elucidation, and biological activity. J Antibiot (Tokyo) 1995, 48(2):119-125.
• [5]Maehr H, Liu CM, Palleroni NJ, Smallheer J, Todaro L, Williams TH, Blount JF: Microbial products. VIII. Azinothricin, a novel hexadepsipeptide antibiotic. J Antibiot (Tokyo) 1986, 39(1):17-25.
• [6]Hayakawa Y, Nakagawa M, Toda Y, Seto H: A new depsipeptide antibiotic, citropeptin. Agric Biol Chem 1990, 54(4):1007-1011.
• [7]Matsumoto N, Momose I, Umekita M, Kinoshita N, Chino M, Iinuma H, Sawa T, Hamada M, Takeuchi T: Diperamycin, a new antimicrobial antibiotic produced by Streptomyces griseoaurantiacus MK393-AF2. I. Taxonomy, fermentation, isolation, physico-chemical properties and biological activities. J Antibiot (Tokyo) 1998, 51(12):1087-1092.
• [8]Maskey RP, Fotso S, Sevvana M, Uson I, Grun-Wollny I, Laatsch H: Kettapeptin: isolation, structure elucidation and activity of a new hexadepsipeptide antibiotic from a terrestrial Streptomyces sp. J Antibiot (Tokyo) 2006, 59(5):309-314.
• [9]Umezawa K, Ikeda Y, Naganawa H, Kondo S: Biosynthesis of the lipophilic side chain in the cyclic hexadepsipeptide antibiotic IC101. J Nat Prod 2002, 65(12):1953-1955.
• [10]Hensens OD, Borris RP, Koupal LR, Caldwell CG, Currie SA, Haidri AA, Homnick CF, Honeycutt SS, Lindenmayer SM, Schwartz CD, et al.: L-156,602, a C5a antagonist with a novel cyclic hexadepsipeptide structure from Streptomyces sp. MA6348. Fermentation, isolation and structure determination. J Antibiot (Tokyo) 1991, 44(2):249-254.
• [11]Uchihata Y, Ando N, Ikeda Y, Kondo S, Hamada M, Umezawa K: Isolation of a novel cyclic hexadepsipeptide pipalamycin from Streptomyces as an apoptosis-inducing agent. J Antibiot (Tokyo) 2002, 55(1):1-5.
• [12]Nakagawa M, Hayakawa Y, Adachi K, Seto H: A new depsipeptide antibiotic, variapeptin. Agric Biol Chem 1990, 54(3):791-794.
• [13]Sakai Y, Yoshida T, Tsujita T, Ochiai K, Agatsuma T, Saitoh Y, Tanaka F, Akiyama T, Akinaga S, Mizukami T: GE3, a novel hexadepsipeptide antitumor antibiotic, produced by Streptomyces sp. I. Taxonomy, production, isolation, physico-chemical properties, and biological activities. J Antibiot (Tokyo) 1997, 50(8):659-664.
• [14]Agatsuma T, Sakai Y, Mizukami T, Saitoh Y: GE3, a novel hexadepsipeptide antitumor antibiotic produced by Streptomyces sp. II. Structure determination. J Antibiot (Tokyo) 1997, 50(8):704-708.
• [15]Tsuji RF, Yamakoshi J, Uramoto M, Koshino H, Saito M, Kikuchi M, Masuda T: Anti-inflammatory effects and specificity of L-156,602: comparison of effects on concanavalin A and zymosan-induced footpad edema, and contact sensitivity response. Immunopharmacology 1995, 29(1):79-87.
• [16]Oelke AJ, France DJ, Hofmann T, Wuitschik G, Ley SV: Piperazic acid-containing natural products: isolation, biological relevance and total synthesis. Nat Prod Rep 2011, 28(8):1445-1471.
• [17]Umezawa K, Nakazawa K, Uchihata Y, Otsuka M: Screening for inducers of apoptosis in apoptosis-resistant human carcinoma cells. Adv Enzyme Regul 1999, 39:145-156.
• [18]Umezawa K, Ikeda Y, Kawase O, Naganawa H, Kondo S: Biosynthesis of polyoxypeptin A: novel amino acid 3-hydroxy-3-methylproline derived from isoleucine. J Chem Soc, Perkin Trans 1 (13):1550-1553.
• [19]Kodani S, Bicz J, Song L, Deeth RJ, Ohnishi Kameyama M, Yoshida M, Ochi K, Challis GL: Structure and biosynthesis of scabichelin, a novel tris-hydroxamate siderophore produced by the plant pathogen Streptomyces scabies 87.22. Org Biomol Chem 2013, 11(28):4686-4694.
• [20]Shen JW, Qin DG, Zhang HW, Yao ZJ: Studies on the synthesis of (2S,3R)-3-hydroxy-3-methylproline via C2-N bond formation. J Org Chem 2003, 68(19):7479-7484.
• [21]Fischbach MA, Walsh CT: Assembly-line enzymology for polyketide and nonribosomal Peptide antibiotics: logic, machinery, and mechanisms. Chem Rev 2006, 106(8):3468-3496.
• [22]Kieser TBM, Chater KF, Butter MJ, Hopwood D: Practical Streptomyces genetics:a laboratory manual. Norwich, UK, Norwich: John Innes Foundation; 2000.
• [23]He Y, Sun Y, Liu T, Zhou X, Bai L, Deng Z: Cloning of separate meilingmycin biosynthesis gene clusters by use of acyltransferase-ketoreductase didomain PCR amplification. Appl Environ Microbiol 2010, 76(10):3283-3292.
• [24]Keatinge-Clay AT: The structures of type I polyketide synthases. Nat Prod Rep 2012, 29(10):1050-1073.
• [25]Yadav G, Gokhale RS, Mohanty D: Computational approach for prediction of domain organization and substrate specificity of modular polyketide synthases. J Mol Biol 2003, 328(2):335-363.
• [26]Smith S, Tsai SC: The type I fatty acid and polyketide synthases: a tale of two megasynthases. Nat Prod Rep 2007, 24(5):1041-1072.
• [27]Tsai SC, Ames BD: Structural enzymology of polyketide synthases. Methods Enzymol 2009, 459:17-47.
• [28]Erb TJ, Berg IA, Brecht V, Muller M, Fuchs G, Alber BE: Synthesis of C5-dicarboxylic acids from C2-units involving crotonyl-CoA carboxylase/reductase: the ethylmalonyl-CoA pathway. Proc Natl Acad Sci USA 2007, 104(25):10631-10636.
• [29]Erb TJ, Brecht V, Fuchs G, Muller M, Alber BE: Carboxylation mechanism and stereochemistry of crotonyl-CoA carboxylase/reductase, a carboxylating enoyl-thioester reductase. Proc Natl Acad Sci USA 2009, 106(22):8871-8876.
• [30]Eustaquio AS, McGlinchey RP, Liu Y, Hazzard C, Beer LL, Florova G, Alhamadsheh MM, Lechner A, Kale AJ, Kobayashi Y, et al.: Biosynthesis of the salinosporamide A polyketide synthase substrate chloroethylmalonyl-coenzyme A from S-adenosyl-L-methionine. Proc Natl Acad Sci USA 2009, 106(30):12295-12300.
• [31]Quade N, Huo L, Rachid S, Heinz DW, Muller R: Unusual carbon fixation gives rise to diverse polyketide extender units. Nat Chem Biol 2012, 8(1):117-124.
• [32]Rachid S, Huo L, Herrmann J, Stadler M, Kopcke B, Bitzer J, Muller R: Mining the cinnabaramide biosynthetic pathway to generate novel proteasome inhibitors. Chembiochem 2011, 12(6):922-931.
• [33]Buntin K, Irschik H, Weissman KJ, Luxenburger E, Blocker H, Muller R: Biosynthesis of thuggacins in myxobacteria: comparative cluster analysis reveals basis for natural product structural diversity. Chem Biol 2010, 17(4):342-356.
• [34]Qu X, Jiang N, Xu F, Shao L, Tang G, Wilkinson B, Liu W: Cloning, sequencing and characterization of the biosynthetic gene cluster of sanglifehrin A, a potent cyclophilin inhibitor. Mol Biosyst 2011, 7(3):852-861.
• [35]Xu Z, Ding L, Hertweck C: A branched extender unit shared between two orthogonal polyketide pathways in an endophyte. Angew Chem Int Ed Engl 2011, 50(20):4667-4670.
• [36]Wilson MC, Nam SJ, Gulder TA, Kauffman CA, Jensen PR, Fenical W, Moore BS: Structure and biosynthesis of the marine streptomycete ansamycin ansalactam A and its distinctive branched chain polyketide extender unit. J Am Chem Soc 2011, 133(6):1971-1977.
• [37]Neumann CS, Jiang W, Heemstra JR Jr, Gontang EA, Kolter R, Walsh CT: Biosynthesis of piperazic acid via N5-hydroxy-ornithine in Kutzneria spp. 744. Chembiochem 2012, 13(7):972-976.
• [38]Fujimori DG, Hrvatin S, Neumann CS, Strieker M, Marahiel MA, Walsh CT: Cloning and characterization of the biosynthetic gene cluster for kutznerides. Proc Natl Acad Sci USA 2007, 104(42):16498-16503.
• [39]Ma J, Wang Z, Huang H, Luo M, Zuo D, Wang B, Sun A, Cheng YQ, Zhang C, Ju J: Biosynthesis of himastatin: assembly line and characterization of three cytochrome P450 enzymes involved in the post-tailoring oxidative steps. Angew Chem Int Ed Engl 2011, 50(34):7797-7802.
• [40]Stachelhaus T, Mootz HD, Marahiel MA: The specificity-conferring code of adenylation domains in nonribosomal peptide synthetases. Chem Biol 1999, 6(8):493-505.
• [41]Challis GL, Ravel J, Townsend CA: Predictive, structure-based model of amino acid recognition by nonribosomal peptide synthetase adenylation domains. Chem Biol 2000, 7(3):211-224.
• [42]Martin JL, McMillan FM: SAM (dependent) I AM: the S-adenosylmethionine-dependent methyltransferase fold. Curr Opin Struct Biol 2002, 12(6):783-793.
• [43]von Dohren H, Keller U, Vater J, Zocher R: Multifunctional Peptide Synthetases. Chem Rev 1997, 97(7):2675-2706.
• [44]Li W, Rokni-Zadeh H, De Vleeschouwer M, Ghequire MG, Sinnaeve D, Xie GL, Rozenski J, Madder A, Martins JC, De Mot R: The antimicrobial compound xantholysin defines a new group of Pseudomonas cyclic lipopeptides. PLoS One 2013, 8(5):e62946.
• [45]Claxton HB, Akey DL, Silver MK, Admiraal SJ, Smith JL: Structure and functional analysis of RifR, the type II thioesterase from the rifamycin biosynthetic pathway. J Biol Chem 2009, 284(8):5021-5029.
• [46]Koglin A, Lohr F, Bernhard F, Rogov VV, Frueh DP, Strieter ER, Mofid MR, Guntert P, Wagner G, Walsh CT, et al.: Structural basis for the selectivity of the external thioesterase of the surfactin synthetase. Nature 2008, 454(7206):907-911.
• [47]Schwarzer D, Mootz HD, Linne U, Marahiel MA: Regeneration of misprimed nonribosomal peptide synthetases by type II thioesterases. Proc Natl Acad Sci USA 2002, 99(22):14083-14088.
• [48]Lin S, Huang T, Shen B: Tailoring enzymes acting on carrier protein-tethered substrates in natural product biosynthesis. Methods Enzymol 2012, 516:321-343.
• [49]Thomas MG, Burkart MD, Walsh CT: Conversion of L-proline to pyrrolyl-2-carboxyl-S-PCP during undecylprodigiosin and pyoluteorin biosynthesis. Chem Biol 2002, 9(2):171-184.
• [50]Pohlmann V, Marahiel MA: Delta-amino group hydroxylation of L-ornithine during coelichelin biosynthesis. Org Biomol Chem 2008, 6(10):1843-1848.
• [51]Jiang W, Cacho RA, Chiou G, Garg NK, Tang Y, Walsh CT: EcdGHK are three tailoring iron oxygenases for amino acid building blocks of the echinocandin scaffold. J Am Chem Soc 2013, 135(11):4457-4466.
• [52]Shimada N, Morimoto K, Naganawa H, Takita T, Hamada M, Maeda K, Takeuchi T, Umezawa H: Antrimycin, a new peptide antibiotic. J Antibiot (Tokyo) 1981, 34(12):1613-1614.
• [53]Umezawa K, Ikeda Y, Uchihata Y, Naganawa H, Kondo S: Chloptosin, an apoptosis-inducing dimeric cyclohexapeptide produced by Streptomyces. J Org Chem 2000, 65(2):459-463.
• [54]Fox KR, Davies H, Adams GR, Portugal J, Waring MJ: Sequence-specific binding of luzopeptin to DNA. Nucleic Acids Res 1988, 16(6):2489-2507.
• [55]Lingham RB, Hsu AH, O'Brien JA, Sigmund JM, Sanchez M, Gagliardi MM, Heimbuch BK, Genilloud O, Martin I, Diez MT, et al.: Quinoxapeptins: novel chromodepsipeptide inhibitors of HIV-1 and HIV-2 reverse transcriptase. I. The producing organism and biological activity. J Antibiot (Tokyo) 1996, 49(3):253-259.
• [56]Huang X, Roemer E, Sattler I, Moellmann U, Christner A, Grabley S: Lydiamycins A-D: cyclodepsipetides with antimycobacterial properties. Angew Chem Int Ed Engl 2006, 45(19):3067-3072.
• [57]Miller ED, Kauffman CA, Jensen PR, Fenical W: Piperazimycins: cytotoxic hexadepsipeptides from a marine-derived bacterium of the genus Streptomyces. J Org Chem 2007, 72(2):323-330.
• [58]Fehr T, Kallen J, Oberer L, Sanglier JJ, Schilling W: Sanglifehrins A, B, C and D, novel cyclophilin-binding compounds isolated from Streptomyces sp. A92-308110. II. Structure elucidation, stereochemistry and physico-chemical properties. J Antibiot (Tokyo) 1999, 52(5):474-479.
• [59]Zhang H, Chen J, Wang H, Xie Y, Ju J, Yan Y: Structural analysis of HmtT and HmtN involved in the tailoring steps of himastatin biosynthesis. FEBS Lett 2013, 587(11):1675-1680.
• [60]Huang T, Wang Y, Yin J, Du Y, Tao M, Xu J, Chen W, Lin S, Deng Z: Identification and characterization of the pyridomycin biosynthetic gene cluster of Streptomyces pyridomyceticus NRRL B-2517. J Biol Chem 2011, 286(23):20648-20657.
• [61]Ishikawa J, Hotta K: FramePlot: a new implementation of the frame analysis for predicting protein-coding regions in bacterial DNA with a high G + C content. FEMS Microbiol Lett 1999, 174(2):251-253.
• [62]Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ: Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 1997, 25(17):3389-3402.
• [63]Ansari MZ, Yadav G, Gokhale RS, Mohanty D: NRPS-PKS: a knowledge-based resource for analysis of NRPS/PKS megasynthases. Nucleic Acids Res 2004, 32(Web Server issue):W405-W413.
• [64]Rausch C, Weber T, Kohlbacher O, Wohlleben W, Huson DH: Specificity prediction of adenylation domains in nonribosomal peptide synthetases (NRPS) using transductive support vector machines (TSVMs). Nucleic Acids Res 2005, 33(18):5799-5808.
• [65]Gust BKT, Chater K: PCR targeting system in Streptomyces coelicolor A3(2). Norwich U K: The John Innes Foundation; 2002.