Labeling proteins with chemical tools is important for examining natural systems, discovering therapeutic agents and developing protein constructs. These methods offer simple but reliable chemistry to the study of peptides and proteins and thus have gained popularity among chemists and biologists. Despite the fact that the number of successful examples has been largely increased over the past decade, there is still an ongoing need for new reagents with better accessibility and reactivity. Diazonium ions are known to selectively react with tyrosine residues for more than a century. But the harsh condition required for diazotization makes it difficult to use this strategy in biological applications. To address this, bench-stable triazabutadienes are made to release diazonium ions upon mild acidification or photoirradiation. Based on our previous study, imidazole N-alkyl substituted triazabutadienes were synthesized and tested for diazonium ion-releasing rates. Surprisingly, the imidazole N-tert-butyl substituted triazabutadiene showed the fastest rate in neutral and basic aqueous solutions. A subsequent NMR study revealed that this rapid release of diazonium ions might be ascribed to the lack of intramolecular π-interactions. In addition, triazabutadienes can be rendered more basic upon photo-isomerization. A water-soluble triazabutadiene was shown to adjust the pH of aqueous solutions. These findings open up new opportunities in protein labeling with unprecedented ease. Moreover, a boronic acid-based photocrosslinker was synthesized to detect protein-protein interactions of glycoproteins. By incorporating benzophenone with a boronic acid and a terminal alkyne, this photocrosslinker is designed to capture the glycoprotein-substrate complex using the combination of photochemistry and bioorthogonal reactions. In conclusion, this dissertation demonstrates progress in developing new probes for protein labeling and protein-protein interactions. These newly developed strategies offer convenient alternatives to those wishing to explore protein activities.

A catalytic (in Brønsted superacid) and a stoichiometric process were developed to synthesise aryl boronic esters with boron cations via electrophilic arene borylation. The treatment of CatBX (Cat = catecholate; X = Cl, Br) with the triethyl salt [Et3Si][closo-CB11H6Br6] in arene solvent gave a transient boron electrophile that reacted as a synthetic equivalent of [CatB]+ in intermolecular electrophilic aromatic borylation at 25 °C. The by-product of the reaction was a strong Brønsted acid that was able to catalyse arene borylation using CatBH at high temperature. This catalytic process furnished aryl boronic esters in high yield with H2 as the only by-product. The use of the robust and weakly coordinating anion [closo-CB11H6Br6]- and the electrophile-resistant catecholborane were crucial for the catalytic process. The reaction mixture of R2BCl (R2 = Cat, Cl4Cat, Cl2), aprotic amine and AlCl3 mainly gave a borenium salt [R2B(amine)][AlCl4] which was in equilibrium with neutral species as revealed by NMR spectroscopy and reactivity studies. This reaction mixture was effective for the regioselective borylation, by electrophilic aromatic substitution, of a range of N-heterocycles, thiophenes and anilines at room temperature. The transterification in situ provided the synthetically useful and more stable pinacol boronate esters in excellent isolated yield. This process displayed remarkable functional-group tolerance for a boron based strong Lewis acid with weak bases (for example -NMe2), ether, and halogen groups all compatible. This process represents a new and inexpensive one-pot direct arene borylation methodology for producing pinacol boronate esters.

The research presented in this thesis focuses on the study of the reaction between boronic acids and diols and its evaluation as a possible "click" reaction, possibly applicable in bioconjugation and drug delivery. A key feature of this reaction is its reversibility at acidic pH, which could allow the release of a diol-containing drug from a bioconjugate in the acidic environment of late endosome/lysosome, possibly after undergoing receptor mediated endocytosis. Over the last two decades various studies have focused on the study of the conjugation of boronic acids to diols using Alizarin Red S as a fluorescence reporter. In this research we have presented an alternative method based on the batochromic shifts of Alizarin Red S absorbance; this method is particularly advantageous in complex systems with an elevated scattering, such as colloidal dispersions or for binding to complexed active compounds. We have therefore demonstrated that this method allows the determination of equilibrium constants between diols (e.g. catecholamines) and boronic acids. We have also demonstrated that the method allows to follow the kinetics of enzymatic reactions involving catechols; in particular, we have focused on cytochrome P450-mediated reactions such as the conversion of estradiol to 2-hydroxyestradiol using CYP1A2, or the demethylation of 3-methoxytyramine to dopamine using CYP2D6. Once we have established a reliable method for following this reaction on low molecular weight compounds, we have applied it to polymeric bioconjugates. Specifically, we have selected hyaluronic acid (HA) as a biocompatible and biodegradable polymeric backbone and produced derivatives containing boronic acids, catechols and dimethylated catechols (as negative controls). The resulting polymers where characterised via UV-Vis, 1H NMR and SLS, also qualitatively evaluating their cytotoxicity and enzymatic degradability. The conjugates with boronic acids showed the lowest cytotoxicity, and the highest degradability. The complexation of HA-boronic derivatives was then studied; using the same library of diols previously used with low molecular weight compounds, evaluating the effect of the presence of the polysaccharidic macromolecular chain.

Molecular recognition at biomembranes is one of the more poorly understood aspects of fundamental research in physical organic chemistry. Our aim was to improve our understanding of the molecular recognition of polysaccharides at biomembranes, in particular developing synthetic lipids that will recognise and report on the presence of glycosaminoglycans (GAG polysaccharides), like heparin and hyaluronic acid. Elevated levels of hyaluronic acid have been implicated in bladder carcinoma and osteoarthritis, and the use of heparin for medical applications is well documented. We synthesised a boronic acid capped lipid that also bore a fluorinated fluorescent reporter group, which could report on multivalent recognition events at bilayer membranes by fluorescent quenching and changes in the lateral distribution of the reporter groups. These preliminary studies showed these boronic acid capped fluorinated lipids gave a fluorescent signal upon interaction with simple mono- and poly- saccharides, albeit with unexpectedly weak binding to these saccharides. To understand and quantify the weaker binding of saccharides to membrane bound boronic acids a series of novel fluorescent and chromogenic lipids were synthesised that bore the reporter group close to the boronic acid. These studies revealed several underlying factors that had important roles in the recognition of oligosaccharides by boronic acid capped lipids. For the first time the effect of the bilayer on saccharide/boronic acid recognition was quantified, with the membrane weakening the interaction 33-fold. We were able to propose a model for the interaction of saccharides for membrane bound boronic acids that explained many of these unexpected observations.We also devised a parallel approach using GAGs to open or close synthetic membrane channels. Using a GAG to switch on the release of an ion or dye would generate a fluorescent signal that amplifies the original recognition event and improves sensitivity for GAGs. Proof-of-principle studies using palladium ions to open dye-transporting channels were successful and these studies were followed by the synthesis of boronic acid-capped cholates. Incorporation of boronic acid-capped cholates into membranes caused changes in the rate of release of alkali metal ions, which caused an enclosed fluorescent dye to give a signal, in the presence or absence of saccharides. These compounds successfully gave a response to the simple saccharide D-fructose but gave no response to other saccharides tested, including various hyaluronic acids. Although we were not able to develop a selective sensor for GAGs, we have developed a model for saccharide/boronic acid interactions that is a valuable addition to the physical organic chemistry of membranes.

The reaction of base-free Chan-Lam coupling was successfully used for functionalization of surface of mesoporous silica gel. Various aromatic, aliphatic, and heterocyclic compounds were immobilized by a copper-catalyzed reaction of corresponding boronic acids with surface amino groups at mild conditions. Obtained functionalized materials were mesoporous although their surface area decreased after immobilization. The reactivity of some surface functional groups was tested in their characteristic reactions.

Indiana University-Purdue University Indianapolis (IUPUI) / The extracellular matrix (ECM) governs many cellular processes through biochemical and mechanical cues. Particularly, the effect ECM mechanical properties on cells fate has been well established over the years. Many hydrogel systems have been used to mimic the dynamic stiffening processes occurring in ECM. However, changes in ECM stiffness does not fully recapitulate the mechanics of native ECM, as viscoelasticity is also a major factor contributing to ECM dynamic property. This thesis describes the design and characterization of an enzyme-crosslinked hydrogel system that is not only capable of being stiffened on demand, but also can be tuned to obtain viscoelasticity. The first objective of this thesis was to utilize horseradish peroxidase (HRP) to crosslink thiol-norbornene hydrogel and use mushroom tyrosinase (MT) to create secondary DOPA-dimer crosslinks that stiffened the hydrogel. The cytocompatibility of HRP-mediated thiol-norbornene gelation and the effect of stiffening on cell fate was evaluated. The second objective of this thesis represented the first step towards developing a hydrogel system whose viscoelasticity could be dynamically tuned. Thiol-norbornene hydrogel was designed to yield dynamically adaptable boronic ester bonds via partial enzymatic reaction. Thiol-norborne hydrogel was made to contain hydroxyl phenol as well as boronic acid residues within its network. MT, in this case was used to oxidize the hydroxy phenol moieties into DOPA, which then complexed with boronic acid, created dynamic bonds, introducing viscoelasticity to an initial elastic hydrogel.

No description available.

Thesis advisor: Jianmin Gao / Besides their broad applications in organic chemistry, boronic acids are also increasingly seen in a variety of biological applications. For instance, they have been used in therapeutic drugs for chemotherapy or probes for the detection of saccharides. One unique feature of boronic acids is that they are capable of forming reversible complexes with sugars or even amino acids. Importantly, they are known to have low inherent toxicity to human. In this work, we have focused on two important reactions involving boronic acids: boronate ester formation, in which boronic acids react with diols and iminoboronate formation in which boronic acids form dative bonds with neighboring amino groups. We have demonstrated the potential of these reactions in bacteria targeting or protein modification. We envisioned that boronic acids could be used as a great warhead to be included in the development of novel antibiotics to counter antibiotic resistance of bacteria, which has emerged to be a serious clinical problem. Different from conventional antibiotics, we decided to utilize reversible covalent chemistries in the design of bacterium binding probes. Inspired by the diol-rich environment on bacterial surface, the first strategy took advantage of the reaction between boronic acid and diols to form boronate esters. We have rationally designed a linear peptide containing five copies of the ‘Wulff-type’ boronic acids or bicyclic amphiphilic peptides with two copies of boronic acids. We examined their capability of binding to E. coli cells or their bactericidal activity against S. aureus. Furthermore, we established a synthetic peptide library (OBTC) incorporating the 2-acetylphenyl boronic acid (2-APBA) warhead to form iminoboronate with the target-lipid II, a precursor for the biosynthesis of peptidoglycan. Although this library failed to generate any potent peptide hits, it provided useful information regarding the development of a synthetic library as well as the screening process. As an extension of the iminoboronate chemistry, thiazolidinoboronate (TzB) attracted our attention for its unique reaction mechanism, superior kinetics and excellent selectivity towards N-terminal cysteine residues. In this work, we have proposed an additional acyl transfer following TzB formation to turn this reaction into an irreversible conjugation. The new reaction inherits the fast kinetics and outstanding selectivity from the TzB chemistry. Excitingly, the product of TzB mediated acyl transfer survived complex conditions such as SDS-PAGE and LC/MS. This reaction was also applied to modify two recombinant proteins with N-terminal cysteine residues or to create a C5C phage library with two distinct modifications. We have further extended the substrate from cysteine to diaminopropionic acid (Dap), serine and tris base. We were delighted to observe imidazolidino boronate (IzB) formation and oxazolidino boronate (OzB) formation, which led to the design of cysteineresponsive probes or peptides that can be spontaneously cyclized in neutral aqueous conditions. / Thesis (PhD) — Boston College, 2020. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Chemistry.

No description available.

<p>The interaction of boronic acid with diols is reversible and pH-dependent. Boronate groups are able to form complex with 1,2- diol or 1,3-diols at pH values above 9. Therefore, the unique property of boronic esters was employed to exploit controllable self-assembly by three independent mechanisms, each of which is independent of the other two. The three interaction mechanisms are 1) electrostatic attraction between positive polymers and negative surfaces. 2) Polyethylene glycol (PEG)—phenolic polymer complex formation, which is one type of hydrogen bonding. 3) Phenylboronate (PBA) binding to polyols.</p> <p>To exploit these interactions, families of water-soluble and bifunctional copolymers containing pairs of non-interacting groups were prepared and characterized. Characterization includes structure, molecular weight, composition, etc. These bifunctional polymers can specifically interact with two other types of polymers/surfaces. Therefore, it provides a possibility to prepare complex assemblies by using multiple polymer/polymer interactions in one step.</p> <p>The utility of multiple, independent interactions was demonstrated by formation of self-assembled multilayer thin films on both silicon wafers and polystyrene latex particles. Moreover, the formation of well-defined nanoparticle aggregates with three different sizes of polystyrene latex particles was studied to extend the application of controllable self-assembly by multiple interactions. The assembly structures of multilayers and latex aggregates were controllable by adjusting the pH and addition of competitive small molecules.</p> <p>In addition to the study of multilayer self-assembly, a new approach for controllable deposition of latex nanoparticles on surfaces was also exploited. Regenerated cellulose films were chemically modified to fabricate the cellulose films bearing surface phenylboronic acid groups (cellulose-PBA). The poly(glycerol monomethacrylate)- stabilized polystyrene (PGMA-PS) latex particles were used to have reversible, pH-dependent adsorption onto the cellulose-PBA by the interaction of boronic acids and diols. Specific adsorption of PGMA-PS onto cellulose-PBA was observed at pH 10.5, whereas the latex particles were removed at pH 4.</p> / Doctor of Engineering (DEng)