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dc.rights.licenseRestricted to current Rensselaer faculty, staff and students. Access inquiries may be directed to the Rensselaer Libraries.
dc.contributorGross, Richard A.
dc.contributorMcGown, Linda Baine
dc.contributorRyu, Chang Yeol
dc.contributorZha, R. Helen
dc.contributor.authorCentore, Robert
dc.date.accessioned2021-11-03T09:19:24Z
dc.date.available2021-11-03T09:19:24Z
dc.date.created2020-08-14T12:22:45Z
dc.date.issued2020-05
dc.identifier.urihttps://hdl.handle.net/20.500.13015/2557
dc.descriptionMay 2020
dc.descriptionSchool of Science
dc.description.abstractIn the first part of this thesis the polymerization of an oligo(Leu) segment from [poly(ethylene glycol) methyl ether]-phenylalanine ethyl ester to form mPEG-b-Phe(Leu)x diblock is described. By taking advantage of natural self-assembly occurring during the aqueous polymerization, a large decrease in the dispersity of the Leu segment in the diblock is observed along with control over the DP of the growing Leu segment. By studying the self-assembly of the reaction product, a mechanism was formulated where in situ co-assembly of mPEG-b-Phe(Leu)x and oligo(Leu)x co-product facilitates precipitation of a β-sheet co-assembly, preventing peptide hydrolysis and further polymerization, and thus providing a means to lower the dispersity and control the DP (DP = 5.1, dispersity < 1.02).
dc.description.abstractGiven the lack of green peptide synthetic methodologies this thesis focuses on the preparation of polymer-peptide conjugates using protease catalyzed peptide synthesis (PCPS). PCPS is capable of forming peptides in buffered aqueous solutions in seconds to a few hours simply by mixing amino acid ethyl ester monomers with a protease. PCPS has been successfully used to synthesize a variety of hydrophobic homopeptides and random copeptides, where precipitation of the hydrophobic product from the reaction buffer facilitated isolation by centrifugation. PCPS has also been applied towards the synthesis of alternating copeptides, and for the synthesis of N-terminus and C-terminus end-capped peptides. Post-polymerization conjugation of PCPS peptides to lipids and poly(ethylene glycol) (PEG) has also been demonstrated. Despite all these advances, the in situ synthesis (where the peptide chain is polymerized from a non-proteinogenic polymer substrate) of polymer-peptide conjugates by PCPS has yet to be explored. This thesis demonstrates the first successful in situ formation of poly(ethylene glycol)-peptide diblocks (PEG-b-peptide) and glycopeptides.
dc.description.abstractIn the second part of this thesis, the polymerization of oligo(Leu) from a glycosylated phenylalanine ethyl ester non-proteinogenic monomer (Glucose-Phe-OEt) to form Glu-Phe(Leu)x is described. Matrix-assisted laser desorption/ionization time-of-flight mass sepectrometery (MALDI-TOF) and nuclear magnetic resonance (NMR) are used to investigate the composition and structure of reaction precipitate. The capability of different proteases to use the non-proteinogenic substrate is investigated along with the effects of substrate concentration and feed ratio (Glucose-Phe-OEt to Leu-OEt). The formation of three different peptide products was observed: the desired Glucose-Phe(Leu)x, deglycosylated glycopeptide Phe(Leu)x, and oligo(Leu)x side-product. A mechanism is described for the formation of the unexpected deglycosylated glycopeptide side-product where post-polymerization protease facilitated deglycosylation occurs. Additionally, future strategies to increase the efficiency of glycopeptide formation during PCPS are described.
dc.description.abstractPolymer-peptide conjugates are unique in combining the dynamic, self-assembling nature of peptides with the structural properties of a synthetic polymer backbone. Furthermore, given the inherent biocompatibility of the peptide segment, by conjugating to biocompatible polymer backbones (e.g. polysachharides) and other FDA approved synthetic polymers (e.g. poly(ethylene glycol), poly(lactic acid), etc.), exciting opportunities exist in the biomedical realm. Indeed, polymer-peptide conjugates are currently used as tissue engineering scaffolds, biosensors, adhesives, and as drug delivery vectors. Yet despite the utility of these materials, current synthetic methods are largely reliant on solid-phase peptide synthesis (SPPS), liquid-phase peptide synthesis (LPPS), N-carboxyanhydride ring opening polymerization (NCA-ROP), recombinant peptide syntheses (RPS), and combinations of these techniques with post-peptide polymerization coupling chemistries (e.g. click chemistry, carbodiimide coupling, etc.). Of these techniques, SPPS and NCA-ROP are currently the most established and utilized. SPPS is capable of synthesizing sequence specific peptides; however, peptide chain growth during SPPS is a cyclic process requiring the use of protection chemistries along with excessive amounts of coupling reagents, amino acids, and wash solvents. On the other hand, NCA-ROP cannot synthesize sequence specific peptides, but is ideal for synthesizing large degree of polymerization (DP) random peptides and block peptides. To make NCA monomers, however, requires the use of toxic phosgene derivatives and strict reaction conditions (dry organic solvent, inert atmosphere, etc.) during the polymerization.
dc.language.isoENG
dc.publisherRensselaer Polytechnic Institute, Troy, NY
dc.relation.ispartofRensselaer Theses and Dissertations Online Collection
dc.subjectChemistry
dc.titleProtease-catalyzed synthesis of peptide conjugates
dc.typeElectronic thesis
dc.typeThesis
dc.digitool.pid180165
dc.digitool.pid180166
dc.digitool.pid180167
dc.rights.holderThis electronic version is a licensed copy owned by Rensselaer Polytechnic Institute, Troy, NY. Copyright of original work retained by author.
dc.description.degreePhD
dc.relation.departmentDept. of Chemistry and Chemical Biology


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