Fitness of Amorphadiene Production Functional Modules and Yeast Chassis

doi:10.7503/cjcu20130339

Abstract

Abstract:

The key to construct artificial yeast cells to produce amorphadiene is the fitness of heterologous functional modules and chassis. The fitness of functional modules and chassis were studied by optimization of vectors, protein expression and promoters in the functional modules. Two functional modules constructed with centromere vector and episomal vector were introduced into two engineered chassis overexpressing key genes(a truncated 3-hydroxyl-3-methylglutaryl-CoA reductase gene tHMGR and farnesyl diphosphate synthase gene ERG20) in mevalonate(MEV) pathway, and the highest amorphadiene production was 11.2 mg/L in the artificial cell with better fitness. Amorphadiene synthase gene ADS and ERG20 were fused to construct fusion enzyme modules, and the fitness with the selected chassis were improved, resulted in a higher amorphadiene production of 17.5 mg/L. With promoter engineering(TDH3p, TEF1p, PGK1p) in the fusion enzyme modules, an artificial cell with a better fitness of functional module and chassis reached a 71.8 mg/L amorphadiene production.

Key words: Amorphadiene, Yeast chassis, Fusion enzyme, Functional module, Fitness

CLC Number:

O629
Q599

TrendMD:

JIA Yun-Jing, ZHAO Juan, DING Ming-Zhu, YUAN Ying-Jin. Fitness of Amorphadiene Production Functional Modules and Yeast Chassis[J]. Chem. J. Chinese Universities, 2013, 34(12): 2765.

References

[1] Liu D., Du J., Zhao G. R., Yuan Y. J., CIESC Journal, 2011, 62(9), 2391—2397(刘夺, 杜瑾, 赵广荣, 元英进. 化工学报, 2011, 62(9), 2391—2397)

[2] Martin V. J., Pitera D. J., Withers S. T., Newman J. D., Keasling J. D., Nat. Biotechnol., 2003, 21(7), 796—802

[3] Babiskin A. H., Smolke C. D., Mol. Syst. Biol., 2011, 7, 471

[4] Partow S., Siewers V., Bjorn S., Nielsen J., Maury J., Yeast, 2010, 27(11), 955—964

[5] Fang F., Salmon K., Shen M. W., Aeling K. A., Ito E., Irwin B., Tran U. P., Hatfield G. W., Da S. N., Sandmeyer S., Yeast, 2011, 28(2), 123—136

[6] Westfall P. J., Pitera D. J., Lenihan J. R., Eng D., Woolard F. X., Regentin R., Horning T., Tsuruta H., Melis D. J., Owens A., Fickes S., Diola D., Benjamin K. R., Keasling J. D., Leavell M. D., McPhee D. J., Renninger N. S., Newman J. D., Paddon C. J., Proc. Natl. Acad. Sci., 2012, 109(3), E111—E118

[7] Ro D. K., Paradise E. M., Ouellet M., Fisher K. J., Newman K. L., Ndungu J. M., Ho K. A., Eachus R. A., Ham T. S., Kirby J., Chang M. C., Withers S. T., Shiba Y., Sarpong R., Keasling J. D., Nature, 2006, 440(7086), 940—943

[8] Paradise E. M., Kirby J., Chan R., Keasling J. D., Biotechnol. Bioeng., 2008, 100(2), 371—378

[9] Albertsen L., Chen Y., Bach L. S., Rattleff S., Maury J., Brix S., Nielsen J., Mortensen U. H., Appl. Environ. Microbiol., 2011, 77(3), 1033—1040

[10] Asadollahi M. A., Maury J., Schalk M., Clark A., Nielsen J., Biotechnol. Bioeng., 2010, 106(1), 86—96

[11] Kong J. Q., Cheng K. D., Wang L. N., Zheng X. D., Dai J. G., Zhu P., Wang W., Acta Pharm. Sin., 2007, 42(12), 1314—1319(孔建强, 程克棣, 王丽娜, 郑晓东, 戴均贵, 朱平, 王伟. 药学学报, 2007, 42(12), 1314—1319)

[12] Engels B., Dahm P., Jennewein S., Metab. Eng., 2008, 10(3/4), 201—206

[13] Rico J., Pardo E., Orejas M., Appl. Environ. Microbiol., 2010, 76(19), 6449—6454

[14] Tokuhiro K., Muramatsu M., Ohto C., Kawaguchi T., Obata S., Muramoto N., Hirai M., Takahashi H., Kondo A., Sakuradani E., Shimizu S., Appl. Environ. Microbiol., 2009, 75(17), 5536—5543

[15] Verwaal R., Wang J., Meijnen J. P., Visser H., Sandmann G., van den Berg J. A., van Ooyen A. J., Appl. Environ. Microbiol., 2007, 73(13), 4342—4350

[16] Kong J. Q., Wang W., Wang L. N., Zheng X. D., Cheng K. D., Zhu P., J. Appl. Microbiol., 2009, 106(3), 941—951

[17] Jackson B. E., Hart-Wells E. A., Matsuda S. P., Org. Lett., 2003, 5(10), 1629—1632

[18] Keasling J. D., Metab. Eng., 2012, 14(3), 189—195

[19] Conrado R. J., Varner J. D., Delisa M. P., Curr. Opin. Biotechnol., 2008, 19(5), 492—499

[20] Jrgensen K., Rasmussen A.V., Morant M., Nielsen A.H., Bjarnholt N., Zagrobelny M., Bak S., Mller B.L., Curr. Opin. Plant Biol., 2005, 8(3), 280—291

[21] Ohto C., Muramatsu M., Obata S., Sakuradani E., Shimizu S., Appl. Microbiol. Biotechnol., 2010, 87(4), 1327—1334

[22] Dai Z., Liu Y., Huang L., Zhang X., Biotechnol. Bioeng., 2012, 109(11), 2845—2853

[23] Zhou Y. J., Gao W., Rong Q., Jin G., Chu H., Liu W., Yang W., Zhu Z., Li G., Zhu G., Huang L., Zhao Z. K., J. Am. Chem. Soc., 2012, 134(6), 3234—3241

[24] Sun J., Shao Z., Zhao H., Nair N., Wen F., Xu J. H., Zhao H., Biotechnol. Bioeng., 2012, 109(8), 2082—2092

[25] Ho S. N., Hunt H. D., Horton R. M., Pullen J. K., Pease L. R., Gene, 1989, 77(1), 51—59

[26] Shao Z., Zhao H., Zhao H., Nucleic Acids Res., 2009, 37(2), e16

[27] Gietz R. D., Woods R. A., Methods Enzymol., 2002, 350, 87—96

[28] Lindahl A. L., Olsson M. E., Mercke P., Tollbom O., Schelin J., Brodelius M., Brodelius P. E., Biotechnol. Lett., 2006, 28(8), 571—580

[29] Dimster-Denk D., Thorsness M. K., Rine J., Mol. Biol. Cell., 1994, 5(6), 655—665

[30] Donald K. A., Hampton R. Y., Fritz I. B., Appl. Environ. Microbiol., 1997, 63(9), 3341—3344

[31] Parent S. A., Fenimore C. M., Bostian K. A., Yeast, 1985, 1(2), 83—138

[32] Mumberg D., Muller R., Funk M., Nucleic Acids Res., 1994, 22(25), 5767—5768