Yates Matthew D, Mickol Rebecca L, Tolsma Joseph S, Beasley Maryssa, Shepard Jamia, Glaven Sarah M
Center for Biomolecular Science and Engineering, US Naval Research Laboratory, Washington, DC, 20375, USA.
Catalent Pharma Solutions, Kansas City, MO, 64137, USA.
Microb Cell Fact. 2024 Dec 19;23(1):336. doi: 10.1186/s12934-024-02617-5.
Biotechnologies that utilize microorganisms as production hosts for lipid synthesis will enable an efficient and sustainable solution to produce lipids, decreasing reliance on traditional routes for production (either petrochemical or plant-derived) and supporting a circular bioeconomy. To realize this goal, continuous biomanufacturing processes must be developed to maximize productivity and minimize costs compared to traditional batch fermentation processes.
Here, we utilized biofilms of the marine bacterium, Marinobacter atlanticus, to produce wax esters from succinate (i.e., a non-sugar feedstock) to determine its potential to serve as a production chassis in a continuous flow, biofilm-based biomanufacturing process. To accomplish this, we evaluated growth as a function of protein concentration and wax ester production from M. atlanticus biofilms in a continuously operated 3-D printed fixed bed bioreactor. We determined that exposing M. atlanticus biofilms to alternating nitrogen-rich (1.8 mM NH) and nitrogen-poor (0 mM NH) conditions in the bioreactor resulted in wax ester production (26 ± 5 mg/L, normalized to reactor volume) at a similar concentration to what is observed from planktonic M. atlanticus cells grown in shake flasks previously in our lab (ca. 25 mg/L cell culture). The wax ester profile was predominated by multiple compounds with 32 carbon chain length (C; 50-60% of the total). Biomass production in the reactor was positively correlated with dilution rate, as indicated by protein concentration (maximum of 1380 ± 110 mg/L at 0.4 min dilution rate) and oxygen uptake rate (maximum of 4 mmol O/L/h at 0.4 min dilution rate) measurements at different flow rates. Further, we determined the baseline succinate consumption rate for M. atlanticus biofilms to be 0.16 ± 0.03 mmol/L/h, which indicated that oxygen is the limiting reactant in the process.
The results presented here are the first step toward demonstrating that M. atlanticus biofilms can be used as the basis for development of a continuous flow wax ester biomanufacturing process from non-sugar feedstocks, which will further enable sustainable lipid production in a future circular bioeconomy.
利用微生物作为脂质合成生产宿主的生物技术将为脂质生产提供一种高效且可持续的解决方案，减少对传统生产途径（石化或植物源）的依赖，并支持循环生物经济。为实现这一目标，必须开发连续生物制造工艺，以与传统分批发酵工艺相比，最大限度地提高生产率并降低成本。
在此，我们利用海洋细菌大西洋海杆菌的生物膜从琥珀酸盐（即非糖类原料）生产蜡酯，以确定其在基于生物膜的连续流生物制造工艺中作为生产底盘的潜力。为此，我们在连续运行的3D打印固定床生物反应器中评估了大西洋海杆菌生物膜的生长与蛋白质浓度以及蜡酯产量之间的关系。我们确定，在生物反应器中使大西洋海杆菌生物膜暴露于交替的富氮（1.8 mM NH）和贫氮（0 mM NH）条件下，会产生蜡酯（26±5 mg/L，以反应器体积归一化），其浓度与我们实验室之前在摇瓶中培养的浮游大西洋海杆菌细胞所观察到的浓度相似（约25 mg/L细胞培养物）。蜡酯谱以多种具有32个碳链长度（C；占总量的50 - 60%）的化合物为主。反应器中的生物量生产与稀释率呈正相关，不同流速下的蛋白质浓度（在0.4分钟稀释率时最高为1380±110 mg/L）和氧气摄取率（在0.4分钟稀释率时最高为4 mmol O/L/h）测量结果表明了这一点。此外，我们确定大西洋海杆菌生物膜的基线琥珀酸盐消耗率为0.16±0.03 mmol/L/h，这表明氧气是该过程中的限制反应物。
此处呈现的结果是迈向证明大西洋海杆菌生物膜可作为从非糖类原料开发连续流蜡酯生物制造工艺基础的第一步，这将在未来的循环生物经济中进一步实现可持续脂质生产。
