The Chemical Synthetic Investigation of Proteins with Sitespecific Lysine Post-Translational Modifications
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Date
2018-04-30
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Abstract
In the recent decade, an increasing amount of protein post-translational
modifications (PTMs) have been discovered, which are important epigenetic markers
widespread on nucleic and cytoplasmic proteins. Lysine (Lys), as the only ε-amino group
containing amino acid, carries many different PTMs, which play important functions in
the regulation of chromatin structure and gene transcription. Methylation and acylation as
the two main groups of Lys modifications are of great significance to the biochemistry
studies. However, the detailed molecular mechanisms and signaling pathways are
remaining elusive, due to the difficulty of site-specific incorporation into target proteins
for molecular biologists.
Therefore, it becomes a challenging but fruitful task for chemical biologists to
prepare sufficient amount of protein of interests (POIs) with PTMs in their homogeneous
states. Based on the noncanonical amino acid (ncAA) incorporation technique, the Liu
group designed a combinatory methodology for the semi-synthesis of protein with sitespecific
Lys PTMs. Mainly two systems have been set up based on the introduction of
different noncanonical amino acids via amble codon suppression followed by certain
bioorthogonal reactions.
This dissertation will begin with a summary of recent PTM functional studies,
followed by a detailed analysis of two ncAA systems for the preparation of POIs with sitespecific
Lys PTMs, N^ε-(4-azidobenzoxycarbonyl)-δ,ε-dehydrolysine (AcdK) for Lys
methylation, and azidonorlucine (AznL) for Lys acylation. The further biological
investigations are highlighted to demonstrate their applications. Another AcdK based
design for the installation of Lys acylation via KAHA ligation strategy will be discussed,
followed by a brief discussion of some other extensions. Last but not the least, we will
talk about a novel visualization strategy for protein fatty acylation modifications by
HaloTag recognition. Optimizations are still needed for this on-going project.
With the combination of ncAA incorporation and bioorthogonal reactions, we can
theoretically incorporate any Lys PTMs with their native structures into target proteins at
specific sites. The protein products we generate can be directly applied for enzymatic
assays in vitro performing similarly as native proteins. Notably, we also provide two
useful functional groups, aldehyde and azido, as very important chemical biology handles
for other protein functionalization purposes.
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Keywords
Protein chemical biology, Post-translational modification, Lysine, noncanonical amino acids