Self-powered enzyme micropumps

Journal name:
Nature Chemistry
Volume:
6,
Pages:
415–422
Year published:
DOI:
doi:10.1038/nchem.1895
Received
Accepted
Published online

Abstract

Non-mechanical nano- and microscale pumps that function without the aid of an external power source and provide precise control over the flow rate in response to specific signals are needed for the development of new autonomous nano- and microscale systems. Here we show that surface-immobilized enzymes that are independent of adenosine triphosphate function as self-powered micropumps in the presence of their respective substrates. In the four cases studied (catalase, lipase, urease and glucose oxidase), the flow is driven by a gradient in fluid density generated by the enzymatic reaction. The pumping velocity increases with increasing substrate concentration and reaction rate. These rechargeable pumps can be triggered by the presence of specific analytes, which enables the design of enzyme-based devices that act both as sensor and pump. Finally, we show proof-of-concept enzyme-powered devices that autonomously deliver small molecules and proteins in response to specific chemical stimuli, including the release of insulin in response to glucose.

At a glance

Figures

  1. Schematic that shows the enzyme pattern on a surface and triggered fluid pumping by enzymatic micropumps.
    Figure 1: Schematic that shows the enzyme pattern on a surface and triggered fluid pumping by enzymatic micropumps.

    a, Au was patterned onto a PEG-coated glass surface using an e-beam evaporator. The patterned surface was functionalized with a quaternary ammonium thiol, which forms a self-assembled monolayer (SAM) on the Au surface. The negatively charged groups on the enzyme bound selectively to the SAM-functionalized Au patterned surface via electrostatic assembly, which resulted in an enzyme pattern on the surface. b, Catalase enzyme immobilized on the Au pattern causes fluid pumping triggered by the presence of both GOx and glucose, which generates hydrogen peroxide in situ.

  2. Fluid pumping velocity in an enzyme-powered micropump as a function of substrate concentration and reaction rate.
    Figure 2: Fluid pumping velocity in an enzyme-powered micropump as a function of substrate concentration and reaction rate.

    a, Pumping velocity in a catalase-powered micropump increases in the presence of its substrate in a reaction-rate-dependent fashion, at substrate concentrations that ranged from 0.001 M to 0.1 M hydrogen peroxide. b, Pumping velocity in a urease-powered micropump increases on increasing the substrate concentration from 0.001 M to 0.75 M urea. c, Pumping velocity in a lipase-powered micropump shows a concentration-dependent increase at substrate concentrations from 0.001 M to 0.5 M 4-nitrophenyl butyrate. d, Pumping velocity in a GOx-powered micropump increases in a substrate concentration- and reaction-rate-dependent manner from 0.001 M to 1 M glucose. The reaction-rate calculations are based on kcat and KM values for enzymes in solution. Error bars represent standard deviations. The means and standard deviations are calculated for 30 tracer particles. The pumping velocities at different substrate concentrations are statistically different (P < 0.01) (see the Supplementary Information).

  3. Temporal and spatial changes in fluid-pumping velocity for catalase-powered micropumps.
    Figure 3: Temporal and spatial changes in fluid-pumping velocity for catalase-powered micropumps.

    a,b, The fluid-pumping velocity in catalase-powered micropumps in the presence of 50 mM hydrogen peroxide was monitored 50–100 µm away from the enzyme pattern as a function of time at intervals of 30 minutes for a total duration of four hours (a), and as a function of distance away from the Au pattern every 1,000 µm for a total distance of 5,000 µm (b). As shown, the pumping velocity decreased over time and distance. Error bars represent standard deviations. The means and standard deviations are calculated for 30 tracer particles. The pumping velocities at different time intervals are statistically different from the pumping velocity at time t = 5 minutes (P < 0.01). The pumping velocities at different distance intervals are statistically different from the pumping velocity at distance d = 100 µm (P < 0.01) (see the Supplementary Information).

  4. Fluid pumping in enzyme micropumps generated by density-driven flows.
    Figure 4: Fluid pumping in enzyme micropumps generated by density-driven flows.

    a, The fluid-pumping velocity monitored in the upright and inverted pump set-ups showed no significant difference for any of the four enzyme micropumps. Error bars represent standard deviations. The means and standard deviations are calculated for 30 tracer particles. The pumping velocities monitored in the upright and inverted pump set-ups are not statistically different (P > 0.01). b, The fluid-pumping velocity monitored in the double-spacer (two × height of chamber, h) set-up showed an approximately seven-fold increase as compared with the single-spacer (h) set-up for three enzyme micropumps. Error bars represent standard deviations. The means and standard deviations are calculated for 30 tracer particles. The pumping velocities monitored in the single- and double-spacer set-ups are statistically different (P < 0.01) (see the Supplementary Information).

  5. Urease-powered stimuli-responsive autonomous release of dye.
    Figure 5: Urease-powered stimuli-responsive autonomous release of dye.

    a, A general schematic that shows the functionalization of enzyme molecules on a positively charged (quaternary-ammonium-terminated) hydrogel, followed by the triggered release of cargo in the presence of the enzyme substrate. b, The concentration of dye (fluorescein) molecules (units of μM) released from urease-anchored hydrogel as a function of time in the presence of different concentrations of urea, monitored using a UV-vis spectrophotometer. The profile shows an increase in the amount of dye released from the hydrogel with increasing urea concentration. The concentration of fluorescein dye molecules released was calculated from the absorbance values by using a calibration curve measured for the dye (Supplementary Fig. 9). The initial absorbance measurement was recorded 30 minutes after substrate (urea) addition.

  6. GOx-powered stimuli-responsive release of insulin.
    Figure 6: GOx-powered stimuli-responsive release of insulin.

    The solution concentration and percentage of insulin molecules released from a GOx-immobilized hydrogel as a function of time in the presence of different concentrations of glucose monitored using a UV-vis spectrophotometer. The profile shows an increase in the amount of insulin released from the hydrogel with increases in glucose concentration in the surrounding solution. The observed behaviour is a direct consequence of the enzymatic reaction-regulated fluid pumping. The concentration of insulin released was calculated using the molar extinction coefficient, ɛ276 = 6,100 M−1 cm−1 (Supplementary Information and Supplementary Tables 5– 8)50. The initial absorbance measurement was recorded ten minutes after substrate (glucose) addition.

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Author information

  1. These authors contributed equally to this work

    • Samudra Sengupta,
    • Debabrata Patra &
    • Isamar Ortiz-Rivera

Affiliations

  1. Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania 16802, USA

    • Samudra Sengupta,
    • Debabrata Patra,
    • Isamar Ortiz-Rivera,
    • Arjun Agrawal,
    • Krishna K. Dey,
    • Thomas E. Mallouk &
    • Ayusman Sen
  2. Institute of Continuous Media Mechanics, Ural Branch of the Russian Academy of Sciences, Perm 614013, Russia

    • Sergey Shklyaev
  3. Department of Chemical Engineering, University of Puerto Rico-Mayagüez, Mayagüez, PR 00681, Puerto Rico

    • Ubaldo Córdova-Figueroa

Contributions

S.S., D.P., T.E.M. and A.S. designed the research. S.S., D.P., I.O-R. and A.A. performed the research. S.S., D.P. and I.O-R. contributed new reagents and analytical tools. S.S., I.O-R., K.K.D., S.Sh., U.C-F., T.E.M. and A.S. analysed the data. S.S., D.P., I.O-R and A.S. wrote the manuscript.

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The authors declare no competing financial interests.

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