Discovery of Targeted Material Binding Microorganisms Using a Centrifugal Microfluidic Platform

Han, Song‐I; Sarkes, Deborah A.; Jahnke, Justin P.; Hurley, Margaret M.; Ugaz, Victor M.; Renberg, Rebecca L.; Sumner, James J.; Stratis‐Cullum, Dimitra N.; Han, Arum

doi:10.1002/admt.202100282

Cited by 1 publication

(21 citation statements)

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“…33−35 Histidine (H) has been flagged as a potential gold binder in our previous work as well as in early attempts to develop design rules for inorganic binding peptides. 24,33 The increase in glycine (G) is also consistent with these design rules, as well as in simulation studies flagging the importance of peptide flexibility in the binding process. 34 Serine (S) has been postulated to play a role in gold binding both experimentally 35 and from simulation, where it has been proposed to facilitate motion to the surface and function as an anchor point.…”

Section: Characterization Of Isolated Single Colonies For Their Mater...supporting

confidence: 75%

“…The capability of the developed magnetophoretic microfluidic device in separating cells having different affinities for gold was assessed using two different Escherichia coli strains (A68 and A3) known to have different affinities toward gold, isolated from a previous work, along with several different negative controls (Table S1). In two separate experiments, cells were injected into the device and then cells flowing out from outlet #1 were collected.…”

Section: Resultsmentioning

confidence: 99%

“…First, surface-binding spot assays were conducted on gold and ITO substrates to test the affinity as well as specificity of the cells. Unless otherwise stated, M6G9, H6G9, and p3-Au12 were used as positive controls against gold (Table S1), and NC as well as NC-P2X used as negative controls (Figure a) . The result of the surface-binding spot assay for all 50 colonies isolated from the final sorting round (round 4) is shown in Figure b.…”

Section: Resultsmentioning

confidence: 99%

“…To analyze the sorted gold-binding isolates, cells from round 1 sorting and round 4 sorting were tested using three complementary methods: a surface-binding spot assay to confirm gold-binding affinity, flow cytometry to monitor P2X expression, and DNA sequencing to determine the displayed peptide sequence. In our previous study, we have observed that the isolated “original” colonies directly obtained from the agar plates after sorting showed a relatively low material binding level as compared to the positive control gold binder strains . Thus, a retransformation process was carried out by isolating each plasmid from the original colonies, followed by transformation of the isolated plasmids into the “Z-competent” MC1061 E.…”

Section: Resultsmentioning

confidence: 99%

“…In addition, measuring the binding strength of microorganisms to the target surface using such microfluidic systems is also quite time-consuming. To overcome these limitations, recently a centrifugal force-based microfluidic biopanning platform was developed . This microfluidic device consists of a simple straight microfluidic channel bound to a glass substrate on which a target material of interest is deposited, and by rotating the device on a spin coater, a centrifugal force-driven flow is generated within the microchannel.…”

Section: Introductionmentioning

confidence: 99%

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Identification of Microorganisms that Bind Specifically to Target Materials of Interest Using a Magnetophoretic Microfluidic Platform

Han

Sarkes

Hurley

et al. 2023

ACS Appl. Mater. Interfaces

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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 materialbinding 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.

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