微生物次級(jí)代謝產(chǎn)物生物合成基因簇與藥物創(chuàng)新

1、 , 150 /FR008/ Secondary metabolic pathway genes and new drug discoveryABSTRACT Microorganisms produce myriads of secondary metabolites with both structural and functional diversities. The cloning of corresponding biosynthetic gene clusters is essential for new drug discovery and yield improvement b

2、y metabolic engineering. To date, more than 150 biosynthetic gene clusters had been cloned via different strategies, which are subsequently manipulated through combinatorial biosynthesis, in vitro glycorandomization, or other biotechnological methods. In our laboratory, several biosynthetic gene clu

3、sters have been cloned and sequenced, representatives of which are responsible for the biosyntheses of the aminoglycoside jinggangmycin/validamycin, polyene antibiotic FR008/candicidin, polyether nanchangmycin, polyketide mEilingmycin, PKSNRPS oxazomycin and others. Extensive analyses of gene functi

4、ons, thEIr unique biosynthetic pathways and regulatory mechanisms have now paved the way for more rational structural modifications through combinatorial biosynthesis and yield improvements using metabolic engineering.KEY WORDS Microbial secondary metabolites; Biosynthetic gene cluster; New drug dis

5、covery; Combinatorial biosynthesis; Metabolic engineering SARS(combinatorial biosynthesis)1(metabolic engineering)2 ()1 1.1 Malpartida19843456 71.2 (3)DNA(4)(5)DNA(8)(9)(10)150(nonribosomal peptide synthaseNRPS)11III(polyketide synthasePKS)111.3 1985Hopwood12 (nonnatural natural product)2001Thorson(

6、in vitro glycorandomizationIVG)13GtfENDP32MalmbergC(LAT)CLATC25142 5102I5102II FR008/HIV 1 2.1 /5102I50084(validamycin)/(validoxylamine)(A)(vogliobose)C7N72515SE50/110acbC500870kb30kb16acbCvalA45kb27(open reading frame,ORF)()1621314(1)A8(valA, B,C,K,L,M,N,G)81326AAValA725ValGAUDPValC AcbMAcbM25()8A1

7、0225102I5102I5102I(),1 / 2.2 FR008/38FR008FR008(candicidin)FR008FR008150kb171498kb21(2)186(modular)fscAF21FR00821(pabABpabC)(mycosamine)(fscMIIfscMIII)(fscMIfscPfscO)FR008FR008FR00818II()fscMIIFscMI 1 2 2 2 FR008/2.3 IPKS908(contig AH)8192021A1325kb30ORF(3)22 11(nanA1A11)1474nanA1

8、A6564kbnan913KRATACPnan10ACPTE (14)CR NanINanONanEnanPnanG54Omethylrhodinose728AAC()3 2.4 avermectin16(4)avermectinC5C432avermectinavermectinavermectinP450aveEaveFaveTE706%659%757%H23avermectinPKS()2.5 (5)(unknown extender unit)ACP(methoxymalonylACP) ()DNA135kb5102II4 avermectinavermectin3 5 1 Hopwo

9、od D A. Genetic contributions to understanding polyketide synthases J. Chem Rev,1997,97(7):24652 Bailey J E, Birnbaum S, Galazzo J L, et al. Strategies and challenges in metabolic engineering J. Ann NY Acad Sci,1990,589:13 Malpartida F, Hopwood D A. Molecular cloning of the whole biosynthetic pathwa

10、y of a Streptomyces antibiotic and its expression in a heterologous host J. Nature,1984,309(5967):4624 Malpartida F, Hallam S E, Kieser H M, et al. Homology between Streptomyces genes coding for synthesis of different polyketides used to clone antibiotic biosynthetic genes J. Nature,1987,325(6107):8

11、185 Cortes J, Haydock S F, Roberts G A, et al. An unusually large multifunctional polypeptide in the erythromycinproducing polyketide synthase of Saccharopolyspora erythraea J. Nature,1990,348(6297):1766 Donadio S, Staver M J, McAlpine J B, et al. Modular organization of genes required for complex p

12、olyketide biosynthesis J. Science,1991,252(5006):6757 Chater K F. The improving prospects for yield increase by genetic engineering in antibioticproducing Streptomycetes J. Biotechnology (N Y),1990,8(2):1158 Kim C G, Yu T W, Fryhle C B, et al. 3Amino5hydroxybenzoic acid synthase, the terminal enzyme

13、 in the formation of the precursor of mC7N units in rifamycin and related antibiotics J. J Biol Chem,1998,273(11):60309 Stratmann A, Mahmud T, Lee S, et al. The AcbC protEin from Actinoplanes species is a C7cyclitol synthase related to 3dehydroquinate synthases and is involved in the biosynthesis of

14、 the alphaglucosidase inhibitor acarbose J. J Biol Chem,1999,274(16):1088910 Piel J, Hertweck C, Shipley P R, et al. Cloning, sequencing and analysis of the enterocin biosynthesis gene cluster from the marine isolate Streptomyces maritimus: evidence for the derailment of an aromatic polyketide synth

15、ase J. Chem Biol,2000,7(12):94311 Marahiel M A, Stachelhaus T, Mootz H D. Modular peptide synthetases involved in nonribosomal peptide synthesis J. Chem Rev,1997,97(7):265112 Hopwood D A, Malpartida F, Kieser H M, et al. Production of hybrid antibiotics by genetic engineering J. Nature,1985,314(6012

16、):64213 Barton W A, Lesniak J, Biggins J B, et al. Structure, mechanism and engineering of a nucleotidylyltransferase as a first step toward glycorandomization J. Nat Struct Biol,2001,8(6):54514 Malmberg L H, Hu W S, Sherman D H. Precursor flux control through targeted chromosomal insertion of the l

17、ysine epsilonaminotransferase (lat) gene in cephamycin C biosynthesis J. J Bacteriol,1993,175(21):691615 Dong H, Mahmud T, Tornus I, et al. Biosynthesis of the validamycins: identification of intermediates in the biosynthesis of validamycin A by Streptomyces hygroscopicus var. limoneus J. J Am Chem

18、Soc,2001,123(12):273316 Yu Y, Bai L, Minagawa K, et al. Gene cluster responsible for validamycin biosynthesis in Streptomyces hygroscopicus subsp. jinggangensis 5008 J. Appl Environ Microbiol,2005,71(9):506617 Hu Z, Bao K, Zhou X, et al. Repeated polyketide synthase modules involved in the biosynthe

19、sis of a heptaene macrolide by Streptomyces sp. FR008 J. Mol Microbiol,1994,14(1):16318 Chen S, Huang X, Zhou X, et al. Organizational and mutational analysis of a complete FR008/candicidin gene cluster encoding a structurally related polyene complex J. Chem Biol,2003,10(11):106519 Sun Y, Zhou X, Liu J, et al. Streptomyces nanchangensis, a producer of the insecticidal polyether