Department of Electrical and Computer Engineering, Texas A&M University, College Station, Texas 77843, USA.
Biotechnology Branch, U.S. Army Combat Capabilities Development Command (DEVCOM), Army Research Laboratory (ARL), Adelphi, Maryland 20783, USA.
ACS Appl Mater Interfaces. 2023 Mar 8;15(9):11391-11402. doi: 10.1021/acsami.2c15192. Epub 2023 Feb 27.
Discovery of microorganisms and their relevant surface peptides that specifically bind to target materials of interest can be achieved through iterative biopanning-based screening of cellular libraries having high diversity. Recently, microfluidics-based biopanning methods have been developed and exploited to overcome the limitations of conventional methods where controlling the shear stress applied to remove cells that do not bind or only weakly bind to target surfaces is difficult and the overall experimental procedure is labor-intensive. Despite the advantages of such microfluidic methods and successful demonstration of their utility, these methods still require several rounds of iterative biopanning. In this work, a magnetophoretic microfluidic biopanning platform was developed to isolate microorganisms that bind to target materials of interest, which is gold in this case. To achieve this, gold-coated magnetic nanobeads, which only attached to microorganisms that exhibit high affinity to gold, were used. The platform was first utilized to screen a bacterial peptide display library, where only the cells with surface peptides that specifically bind to gold could be isolated by the high-gradient magnetic field generated within the microchannel, resulting in enrichment and isolation of many isolates with high affinity and high specificity toward gold even after only a single round of separation. The amino acid profile of the resulting isolates was analyzed to provide a better understanding of the distinctive attributes of peptides that contribute to their specific material-binding capabilities. Next, the microfluidic system was utilized to screen soil microbes, a rich source of extremely diverse microorganisms, successfully isolating many naturally occurring microorganisms that show strong and specific binding to gold. The results show that the developed microfluidic platform is a powerful screening tool for identifying microorganisms that specifically bind to a target material surface of interest, which can greatly accelerate the development of new peptide-driven biological materials and hybrid organic-inorganic materials.
通过对具有高多样性的细胞文库进行迭代的基于生物淘选的筛选，可以发现特异性结合目标材料的微生物及其相关表面肽。最近，已经开发并利用基于微流控的生物淘选方法来克服传统方法的局限性，在传统方法中，控制施加的剪切力以去除不结合或仅弱结合到目标表面的细胞是困难的，并且整个实验过程是劳动密集型的。尽管这些微流控方法具有优势并且成功地证明了它们的实用性，但这些方法仍需要几轮迭代的生物淘选。在这项工作中，开发了一种基于磁泳的微流控生物淘选平台来分离与目标材料（在本例中为金）结合的微生物。为了实现这一目标，使用了仅附着在对金表现出高亲和力的微生物上的金涂覆的磁性纳米珠。该平台首先用于筛选细菌肽展示文库，其中仅可以通过在微通道内产生的高梯度磁场分离到特异性结合金的细胞，从而即使在仅一轮分离后，也可以对具有高亲和力和特异性的许多分离株进行富集和分离。对分离出的微生物的氨基酸组成进行分析，以更好地了解对其特异性结合材料的能力有贡献的肽的独特属性。接下来，利用微流系统筛选土壤微生物，这是丰富的极端多样化微生物的来源，成功地分离出许多对金表现出强烈和特异性结合的天然存在的微生物。结果表明，所开发的微流控平台是一种强大的筛选工具，可用于鉴定特异性结合目标材料表面的微生物，这可以极大地加速新型肽驱动的生物材料和混合有机-无机材料的开发。
