Among all 20 native amino acids in proteins, lysine (Lys) undergoes the most diverse forms of posttranslational modifications (PTMs). The unique nucleophilicity of its side chain amine allows Lys to be selectively modified with several types of alkylation and a number of small-molecule and protein acylations. These PTMs, especially in eukaryotes, regulate enzyme activities, interactions of proteins with their partners, cellular localization of proteins, and protein metabolism. Abnormality of these PTMs correlates with the development of many diseases. Although important, biochemical studies of Lys PTMs are cumbersome due to the difficulty to synthesize proteins with them. Several methods have been developed for the synthesis of proteins with Lys PTMs. However, they cannot be generally applied. With an overall objective to formulate straightforward methods that can be generally applied for the synthesis of proteins with Lys PTMs, the current application will focus on the development of amber suppression-based noncanonical amino acid (ncAA) mutagenesis methods in combination with chemical transformation for the installation of Lys PTMs into proteins.
Three specific aims will be pursued: 1) Develop enhanced amber suppression-based ncAA mutagenesis methods for the recombinant synthesis of proteins with Lys alkylations and small-molecule acylations; 2) Develop amber suppression-based ncAA mutagenesis methods for the synthesis of proteins with ubiquitin and ubiquitin-like protein modifications; and 3) Formulate recombinant strategies in conjunction with biocompatible reactions to synthesize proteins installed site-specifically with two different Lys PTMs. The successful completion of the proposed study will make available straightforward methods for the synthesis of proteins with most Lys PTMs for their functional investigation.

Public Health Relevance

The proposed research is relevant to public health because it aims to develop methods for the synthesis of proteins with lysine posttranslational modifications for understanding human health and diseases. The project is relevant to NIH?s mission in promoting health and combating diseases because it will assist our understanding of posttranslational lysine modifications in cell development and differentiation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM127575-03
Application #
10063529
Study Section
Synthetic and Biological Chemistry B Study Section (SBCB)
Program Officer
Bond, Michelle Rueffer
Project Start
2019-01-15
Project End
2022-11-30
Budget Start
2020-12-01
Budget End
2021-11-30
Support Year
3
Fiscal Year
2021
Total Cost
Indirect Cost
Name
Texas A&M University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
020271826
City
College Station
State
TX
Country
United States
Zip Code
77845