Exploring the MOF-peptide interface: from phage display to materials synthesis, thin films and composites
Lead Research Organisation:
Nottingham Trent University
Department Name: School of Science & Technology
Abstract
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| Description | The key findings of this project are summarised below: The combinatorial phage display (Ph.D.) method can be used to identify strongly-binding peptide sequences for metal-organic frameworks (MOFs), as demonstrated for a number of common MOF materials including UiO-66, ZIF-8, HKUST-1 and Ti-MIL-125. Full sequence analysis supported by simulation approaches allow us to identify which specific amino-acid residues or combination of residues are most recurring and important to MOF binding, which may permit future targeted peptide design. In collaboration with the team of Hendrik Heinz at Boulder, we have developed simulation approaches to investigate the binding mechanism of peptides toward MOFs. This has focused on the UiO-66-X system and both on- and off-target binding are considered. The simulations have been highly successful in reproducing the order of binding observed during Ph.D., allowing us to visualize which MOF-peptide interactions are important and effective modelling of the surface including the interactions of water molecules with the amino-functional groups in UiO-66-NH2. Further, the Heinz group, in developing a specific forcefield for these systems have show the instability of particular crystal faces, with loss of Zirconium ions and hydroxide ions from surfaces in a plane specific fashion that helps us to understand the zeta potential data where materials give positive zeta potential values. Framework functionalisation can lead to a unique-binding sequence: amino-functionalization of UiO-66 results in a unique binding sequence being identified from a phage library of 10^9 possible sequences whereas the non-amino derivative yields a smaller number of binding sequences. This is verified using non-functionalized ZIF-8, and implies that frameworks bearing additional functional groups have a greater potential to interact strongly with phage-bound peptides. Different sequences have also been identified when the framework crystallinity, topology or crystal morphology is changed but composition remains constant such as in the polymorphic ZIF system. In this latter case, two sequences are identified, one which binds to all polymorphs investigated (a general-binder) and one that is specific to sodalite. We have also observed some degree of discrimination between sodalite crystals of differing morphology, which suggests that the Ph.D. peptides are able to bind to the MOF in a facet-selective manner. This may open the door to highly controlled surface functionalisation of MOF crystals in the future. The binding of peptide sequences identified by Ph.D. to MOFs can be determined using isothermal calorimetry (ITC) and changes in zeta-potential (ZP) on peptide-binding. Typical dissociation constants are in the range 10-6 M-1, and it is possible to extract thermodynamic properties such as ?G values from the ITC data. A complete study was performed on UiO-66 and the amino-functionalised analogue UiO-66-NH2 and while peptide-binding was found to be enthalpically-driven the differences between on- and off-target MOF-peptide binding were more similar than expected. This is attributed to the differences in peptide behaviour between the Ph.D. screening and the binding tests in solution and the high degree of aggregation of the relatively hydrophobic peptides when in solution. The experimental binding observations have been consistently reproduced using simulation in most cases. Binding studies are further supported by synchrotron radiation circular dichroism (SRCD) spectroscopy, applied to study the MOF-peptide interface in this project for the first time. By comparison of the peptides alone in solution and in the presence of the desired MOF (UiO-66-X or ZIFs) then conformational changes upon binding can be monitored. This is further supported by computational approaches, where simulated Ramachandran plots of the torsional angle landscape significantly aids interpretation of the experimental data. Overall this has been an excellent addition to the toolbox of methods to study how peptides interact with MOFs. An extensive study of the binding of titania-binding peptides toward Ti-MIL-125 has also been undertaken as part of this work. We aimed at a comparison of binding affinities of the MOF PH.D. sequences against the two MOFs in comparison to anatase and rutile. Additionally, we started a study with three well-known Titania binding peptides (Ti-1, Ti-2, Ti-1231) as references sequences and compare their binding affinity against anatase, rutile, Ti-Mil-125 and Ti-MIL-125-NH2. In some cases we find that the titania peptides also bind strongly to the MOFs, perhaps indicating structural similarity between the minerals and the inorganic MOF secondary-building unit clusters. Again, SRCD revealed that we find the strongest pronounced conformational changes between titania-peptides and their interaction with Ti-Mil-125-NH2. The atomistic interpretation is however complex due to the strong self-agglomeration behavior of the investigated peptides in solution, which was further confirmed by ITC and dynamic light scattering experiments. Addition of Ph.D. peptides to MOF syntheses in a biomimetic mineralisation approach has been found to influence crystallinity and particle morphology in the UiO-66-X system, when an aqueous synthesis is employed. Interestingly, the peptides can guide framework assembly under modified conditions where this otherwise forms only amorphous products, directly mirroring natural biomineralisation approaches. Control experiments with just the amino acids or under conditions where the Ph.D. peptides fragment do not yield crystalline material indicating there is clear added value in using the peptides for MOF synthesis and that the effect of pH is minimal. Addition of the Ph.D. peptides to the ZIF-8 synthesis, especially when starting from a point that otherwise yields an amorphous project, has the capacity to influence framework topology, and in some cases can increase the stability and persistence of metastable phases so can also modulate the kinetics of framework assembly. This follows the order of frequency in which the peptides were found during phage display screening, and general-binding peptides allow phase transformation between ZIF polymorphs but where a sequence was specifically identified for a single topology this is significantly stabilized vs. the control. In one case, we observed the recently reported katsenite phase, a topology that has only previously been accessed through ball-milling of the precursors, which further demonstrates the power of this biomimetic approach. As expected the best biomimetic framework assembly results are obtained under mild reaction conditions where the peptide is demonstrably intact. We have found that biologically benign Ca-MOFs can be prepared directly from a range of calcium carbonate polymorphs in high yield in water under ambient conditions. Peptides previously found by Ph.D. to bind strongly to calcium carbonate are currently under investigation to determine if they can influence framework topology, morphology or crystallinity. Control over Ca-MOFs synthesis and shape is directly relevant to the use of MOFs in biomedicine, and given the prevalence of CaCO3 in natural systems provides an excellent start point to further investigate biomimetic MOF synthesis approaches. Overall the above findings clearly demonstrate that the application of genetically modified peptide libraries can identify strong binding sequences for MOFs, and that use of these peptides in biomimetic mineralisation can direct the topology, modulate assembly kinetics and influence the crystallinity of these important framework materials in a predictable manner. This is important, as it demonstrates that small biological units can also direct materials assembly and that the mechanisms of action go beyond simple charge effects as reported for MOF-protein systems. The ability to use peptides and identify specific residues of importance through screening and simulation offers new vistas in biomimetic MOF synthesis and an increased understanding of how this important class of material interacts with biological building blocks relevant to their use in drug delivery and bio-applications more widely. Further specific details in respect of the binding of phage identified peptides to the ZIF-8 series of MOFs, titanium containing MOFs and HKUST-1 are given below. Zif-8 series: We identified previously two peptides (P1, P2) via phage display against four (pseudo)polymorphs: Zif-8, Zif-8-truncated, ZiP and Zif-L. To study the binding affinities, we used UV-vis adsorption isotherms and synchrotron radiation circular dichroism (SRCD) spectroscopy for conformational changes upon interaction. P1 shows, except for Zif-8 a concentration depended binding behavior. For this reason, we used a dose-response (sigmoidal) fitting approach for the interaction of P1 with Zif-8-t, ZiP and Zif-L. For the interaction with Zif-8 we utilized the Hill adsorption isotherm to obtain the binding affinities. We find the highest affinity of P1 for Zif-8 with ?G=7.5 kcal mol-1and very similar values for the other (pseudo) polymorphs (around ?G=6.7 kcal mol-1). Generally, the concentration dependent binding behavior appears be in good agreement with the biomineralization results. Based on our SRCD spectroscopy study P1 shows an unordered conformation in solution and consistent conformational changes for the investigated Zif series. There changes indicate an increase in order upon adsorption and additionally similar binding configurations, more precisely conformational ensemble. P2 binding to all (pseudo) polymorphs can consistently be fitted with the Hill adsorption isotherm. The strongest interaction can be found for Zif-8 (?G=7.3 kcal mol-1) and Zif-8-t (?G=7.2 kcal mol-1) in agreement with their origin in phage display. We find only a very weak interaction of the peptide with Zif-L (?G=4.5 kcal mol-1) additionally the conformation remains unchanged indicating the absence of interaction. For the other (pseudo)polymorphs we find again consistent changes. Titanium containing MOFs: We identified previously two peptides (P1, P3) via phage display against Ti-Mil-125 and Ti-MIL-125-NH2. We aim at a comparison of binding affinities of this sequences against the two MOFs in comparison to anatase and rutile. Additionally, we started a study with three well-known Titania binding peptides (Ti-1, Ti-2, Ti-1231) as references sequences and compare their binding affinity against anatase, rutile, Ti-Mil-125 and Ti-MIL-125-NH2. For the determination of ?G we isothermal titration calorimetry, which additionally allows for the determination of ?H and T?S. Preliminary results indicate that Ti-1 has the highest affinity towards rutile (?G=9.7 kcal mol-1), while the binding towards Ti-Mil-125-NH2 is significantly reduced (?G=7.6 kcal mol-1). Ti-2 shows in comparison a higher affinity towards Ti-Mil-125-NH2 (?G=9.2 kcal mol-1) than to anatase and rutile, ?G=8.3 kcal mol-1 and ?G=7.8 kcal mol-1 respectively. Ti-1231 shows the highest affinity towards rutile (?G=10.0 kcal mol-1) and the same values for anatase and Ti-Mil-125-NH2 (?G=9.2 kcal mol-1). SRCD revealed that we find the strongest pronounced conformational changes of Ti-1, Ti-2 and Ti-1231 for the interaction with Ti-Mil-125-NH2. The atomistic interpretation of is however complex due to the strong self-agglomeration behavior of the investigated peptides in solution, which we confirmed by ITC and dynamic light scattering experiments. HKUST-1: We identified previously two peptides (P1 and P4) via phage display against a selected Cu-based MOF (HKUST-1). We utilize this two peptides for optimization of surface-anchored MOFs (SURMOFs) deposition. For the preparation of the SURMOF we compare two different approaches: a dipping method and a surface functionalization using the setup of a surface plasmon resonance (SPR) spectrometer. We used the dipping approach to optimize the conditions (solvent, incubation, washing, etc.) found in literature to achieve a higher surface coverage and homogeneity after 50 dipping cycles. First tests towards green synthesis conditions revealed the lack of suitability of aqueous conditions. The knowledge obtained from dipping method was used to adapt conditions for the deposition of HKUST-1 on Au sensors within the SPR setup. This approach will allow for the characterization of the kinetics during the step-by-step assembly of the SURMOF. Preliminary studies suggest that the addition of peptides prior the assembly of HKUST further enhances the crystallinity and surface coverage. The assembly will be characterized using Fourier Transform Infrared Spectroscopy (FT-IR) and Scanning Electron Microscopy (SEM) together with the SPR sensograms to understand the synthesis, reaction kinetics and thermodynamics. Unfortunately, this approach can only be used for up to 10 layers as beyond this the surface build up is not detectable using SPR. |
| Exploitation Route | The development of bioinspired routes to materials including other MOFS. Generation of materials that can be used in sensing and biotechnology. |
| Sectors | Chemicals,Pharmaceuticals and Medical Biotechnology |
| Description | The impact of this work is recorded against grant ref EP/N025822/1 |
| First Year Of Impact | 2023 |
| Description | Probing mechanisms of biological-material interactions; towards realizing biominetic materials by understanding molecular-level interactions |
| Amount | $510,000 (USD) |
| Funding ID | FA9550-16-1-0213 |
| Organisation | Airforce Office of Scientific Research |
| Sector | Public |
| Country | United States |
| Start | 05/2016 |
| End | 05/2019 |
| Description | Simulations for MOFS |
| Organisation | University of Colorado Boulder |
| Country | United States |
| Sector | Academic/University |
| PI Contribution | Experimental data to allow development of forcefield for specific MOFS explored in the project. Experimental data on peptide identification from phage display, binding data and biomineralization data. |
| Collaborator Contribution | Development of new forcefield for UiO66 and UiO66-NH2. Development of understanding of surface stability of different crystal faces. Probe of peptide/surface interactions- generation of data to support experimental data on binding and MOF behaviour collected in my laboratory. |
| Impact | Not yet available |
| Start Year | 2018 |
| Description | Keynote research talk |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Other audiences |
| Results and Impact | keynote research lecture- TMS meeting, San DIego- dissemination- may influence a scientists from Spain to spend sabbatical in our lab. |
| Year(s) Of Engagement Activity | 2020 |
| Description | MRS Fall 2018 Invited talk |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Other audiences |
| Results and Impact | Invited lecture at MRS meeting, Boston USA where we described our research at the abiotic: biotic interface. |
| Year(s) Of Engagement Activity | 2018 |
| Description | Research talk during collaboration visit to Boulder, Colorado |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Postgraduate students |
| Results and Impact | Research talk at University of Boulder during a visit to our collaborator, Dr Heinz. |
| Year(s) Of Engagement Activity | 2020 |
| Description | Sitges Bio meeting March 2019 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Postgraduate students |
| Results and Impact | Contributed research talk |
| Year(s) Of Engagement Activity | 2019 |
| Description | Snow and Science talk- Dr Monika Michaelis |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Postgraduate students |
| Results and Impact | communicating science between research groups involved in computayional research. |
| Year(s) Of Engagement Activity | 2020 |