Synthesis of Epidermal Growth Factor by Native Chemical Ligation and Incorporation of Non-Natural Amino Acids
type: CreativeWork
description: The objective of this study was to develop a method for the efficient and rapid generation of analogs of hEGF and to incorporate unnatural amino acids into EGF and its fragments. Since the analogs share sequences, for optimal versatility and economy, it was decided that hEGF would be assembled from fragments which could be used as building blocks in the synthesis of several analogs. Coupling of amino acids to the conformationally constrained residues Tic and HOTic, is difficult. In model experiments, use of HOAt in combination with either DIC or F1ATU for activation was effective in solving these coupling difficulties. HOTic was then incorporated into the 20-31 fragment of hEGF. [Abu'ui,HOTic ;]hEGF(20-31 )-NH- was shown to be a 'difficult sequence', but replacement of Tyr at position 29 with HOTic facilitates the complete dodecapeptide synthesis. Chemoselective ligation techniques allow the use of unprotected peptide fragments which can eliminate solubility problems associated with fragment condensation. Native chemical ligation results in a peptide bond thus, joining the fragments. Therefore, it was selected as the method for the synthesis of hEGF. To compromise between yield and flexibility, it was decided that the hEGF would be assembled from three fragments. During the synthesis of the thioester fragments, [Cys (Aem)6]hEGF(l-13)-MPAL and [Cys(Acm)20]hEGF(14-30)-MPAL, significant aspartimide formation occurred. To suppress this side reaction, several methods were examined. 'Low-high' TFMSA cleavage, deprotection with 'low' TFMSA cleavage conditions followed by thiolyzis and use of 2-Ada or i-Bu protection on Asp did not solve the problem. cHx protection on Asp, in combination with HF cleavage, supressed the side reaction, but in the case of Msc-[Cys(Acm)20]hEGF(14-30)-MPAL, further variation of HF cleavage conditions is necessary to eliminate other side reactions. Fmoc-Cys(Phacm)-OH was synthesized and side products were identified. The synthesis of [Cys(Phacm)3…
abstract: The objective of this study was to develop a method for the efficient and rapid generation of analogs of hEGF and to incorporate unnatural amino acids into EGF and its fragments. Since the analogs share sequences, for optimal versatility and economy, it was decided that hEGF would be assembled from fragments which could be used as building blocks in the synthesis of several analogs. Coupling of amino acids to the conformationally constrained residues Tic and HOTic, is difficult. In model experiments, use of HOAt in combination with either DIC or F1ATU for activation was effective in solving these coupling difficulties. HOTic was then incorporated into the 20-31 fragment of hEGF. [Abu'ui,HOTic ;]hEGF(20-31 )-NH- was shown to be a 'difficult sequence', but replacement of Tyr at position 29 with HOTic facilitates the complete dodecapeptide synthesis. Chemoselective ligation techniques allow the use of unprotected peptide fragments which can eliminate solubility problems associated with fragment condensation. Native chemical ligation results in a peptide bond thus, joining the fragments. Therefore, it was selected as the method for the synthesis of hEGF. To compromise between yield and flexibility, it was decided that the hEGF would be assembled from three fragments. During the synthesis of the thioester fragments, [Cys (Aem)6]hEGF(l-13)-MPAL and [Cys(Acm)20]hEGF(14-30)-MPAL, significant aspartimide formation occurred. To suppress this side reaction, several methods were examined. 'Low-high' TFMSA cleavage, deprotection with 'low' TFMSA cleavage conditions followed by thiolyzis and use of 2-Ada or i-Bu protection on Asp did not solve the problem. cHx protection on Asp, in combination with HF cleavage, supressed the side reaction, but in the case of Msc-[Cys(Acm)20]hEGF(14-30)-MPAL, further variation of HF cleavage conditions is necessary to eliminate other side reactions. Fmoc-Cys(Phacm)-OH was synthesized and side products were identified. The synthesis of [Cys(Phacm)3…
author: Bozsö, Zsolt
language: en_US
date published: 2004
date modified: 2026-07-24T01:50:46Z
provider: Creighton University
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: Creighton University

PropertyValue
property ID: Handle
value: http://hdl.handle.net/10504/47224