L-酪氨酸是一种重要的营养必需氨基酸,广泛应用于食品、医药、化妆品等行业。非磷酸转移酶葡萄糖转运途径是细菌吸收转运葡萄糖的重要途径,对细菌的新陈代谢具有显著的影响。以解淀粉芽胞杆菌HZΔptsH为出发菌株,表达了不同物种来源的葡萄糖转运蛋白编码基因,探究非磷酸转移酶葡萄糖转运途径基因对解淀粉芽胞杆菌合成L-酪氨酸的影响。摇瓶发酵结果表明,HZΔptsH强化表达自身来源葡萄糖转运蛋白编码基因glcP后,L-酪氨酸产量提高了22%,而表达大肠杆菌的galP基因和谷氨酸棒状杆菌的idolT1基因对L-酪氨酸产量没有显著性的影响。研究证实,过表达解淀粉芽胞杆菌自身glcP基因是一种有效的L-酪氨酸强化策略,该研究为后续解淀粉芽胞杆菌合成L-酪氨酸的代谢工程育种提供了借鉴。
As an important nutritional essential amino acid,L-tyrosine was widely used in food,medical,cosmetics and other industries. The non-phosphotransferase glucose transport pathway was a key way for bacteria to absorb glucose,which had significant effect on the metabolism of bacteria. In this study,the Bacillus amyloliquefaciens HZΔptsH was used as the original strain. Different genes of glucose transporters were expressed to strengthen the glucose absorption transport,and their effects on the biosynthesis of L-tyrosine were investigated. Flasks fermentation results showed that the yield of L-tyrosine was improved by 22% after expression of the native glucose transporter gene glcP in HZΔptsH. Wherein,overexpressing the galP gene from Escherichia coli and idolT1 gene from Corynebacterium glutamicum showed no significant effect on L-tyrosine production. This study confirmed that overexpressing the native glcP gene of B. amyloliquefaciens was an effective strategy for enhancing L-tyrosine production,which provided the reference for further metabolic engineering breeding of B. amyloliquefaciens for L-tyrosine synthesis.
[1] 姚元锋. L-酪氨酸代谢平台构建及其在丹参素合成中的应用[D].天津:天津大学,2013.
[2] DENG W,WANG Y,ZHANG S,et al.Catalytic amino acid production from biomass-derived intermediates[J]. Proceedings of the National Academy of Sciences,2018,115(20): 5 093-5 098.
[3] JANG S,GANG H,KIM B G,et al.FCS and ECH dependent production of phenolic aldehyde and melanin pigment from L-tyrosine in Escherichia coli[J]. Enzyme and Microbial Technology,2018,112: 59-64.
[4] WANG J,GULERIA S,KOFFAS M A G,et al. Microbial production of value-added nutraceuticals[J]. Current Opinion in Biotechnology,2016,37: 97-104.
[5] D′ESTE M,ALVARADO-MORALES M,ANGELIDAKI I. Amino acids production focusing on fermentation technologies-A review[J]. Biotechnology Advances,2018,36(1): 14-25.
[6] XU Y,LI Y,ZHANG L,et al.Unraveling the specific regulation of the shikimate pathway for tyrosine accumulation in Bacillus licheniformis[J]. Journal of industrial Microbiology & Biotechnology,2019,46(8): 1 047-1 059.
[7] RODRIGUEZ A,MARTNEZ J A,FLORES N,et al.Engineering Escherichia coli to overproduce aromatic amino acids and derived compounds[J]. Microbial Cell Factories,2014,13(1): 126.
[8] NODA S,KONDO A.Recent advances in microbial production of aromatic chemicals and derivatives[J]. Trends in Biotechnology,2017,35(8): 785-796.
[9] BREY L F,WODARCZYK A J,THØFNER J F B,et al. Metabolic engineering of Synechocystis sp. PCC 6803 for the production of aromatic amino acids and derived phenylpropanoids[J]. Metabolic Engineering,2020,57: 129-139.
[10] IKEDA M,OKAMOTO K,KATSUMATA R.Cloning of the transketolase gene and the effect of its dosage on aromatic amino acid production in Corynebacterium glutamicum[J]. Applied Microbiology and Biotechnology,1999,51(2): 201-206.
[11] SANTOS C N S,STEPHANOPOULOS G. Melanin-based high-throughput screen for L-tyrosine production in Escherichia coli[J]. Applied and Environmontal Microbiology,2008,74(4): 1 190-1 197.
[12] MUOZ A J,HERNNDEZ-CHVEZ G,DE ANDA R,et al.Metabolic engineering of Escherichia coli for improving L-3,4-dihydroxyphenylalanine (L-DOPA) synthesis from glucose[J]. Journal of Industrial Microbiology & Biotechnology,2011,38(11): 1 845.
[13] FREITAS M A,MEDEIROS F H V,MELO I S,et al. Stem inoculation with bacterial strains Bacillus amyloliquefaciens (GB03) and Microbacterium imperiale (MAIIF2a) mitigates Fusarium root rot in cassava[J]. Phytoparasitica,2019,47(1): 135-142.
[14] TORRES M J,BRANDAN C P,SABATÉ D C,et al.Biological activity of the lipopeptide-producing Bacillus amyloliquefaciens PGPBacCA1 on common bean Phaseolus vulgaris L. pathogens[J]. Biological Control,2017,105:93-99.
[15] LEE J Y,SHIM J M,YAO Z,et al.Antimicrobial activity of Bacillus amyloliquefaciens EMD17 isolated from Cheonggukjang and potential use as a starter for fermented soy foods[J]. Food Science and Biotechnology,2016,25(2): 525-532.
[16] BEZ J L,BOLVAR F,GOSSET G.Determination of 3-deoxy-D-arabino-heptulosonate 7-phosphate productivity and yield from glucose in Escherichia coli devoid of the glucose phosphotransferase transport system[J]. Biotechnology and Bioengineering,2001,73(6): 530-535.
[17] ESCALANTE A,CALDERN R,VALDIVIA A,et al.Metabolic engineering for the production of shikimic acid in an evolved Escherichia coli strain lacking the phosphoenolpyruvate: Carbohydrate phosphotransferase system[J]. Microbial Cell Factories,2010,9(1): 21.
[18] LINDNER S N,SEIBOLD G M,HENRICH A,et al.Phosphotransferase system-independent glucose utilization in Corynebacterium glutamicum by inositol permeases and glucokinases[J]. Applied and Environmental Microbiology,2011,77(11): 3 571-3 581.
[19] IKEDA M,MIZUNO Y,AWANE S,et al.Identification and application of a different glucose uptake system that functions as an alternative to the phosphotransferase system in Corynebacterium glutamicum[J]. Applied Microbiology and Biotechnology,2011,90(4): 1 443.
[20] CHANDRAN S S,YI J,DRATHS K M,et al.Phosphoenolpyruvate availability and the biosynthesis of shikimic acid[J]. Biotechnology Progress,2003,19(3): 808-814.
[21] 赖联贺. 改造谷氨酸棒杆菌非磷酸转移酶葡萄糖转运途径高产L-丝氨酸[D].无锡:江南大学,2017.