Peptide natural products represent a diverse class of important therapeutics. The biosynthetic enzymes responsible for constructing nonribosomal peptides have complex structural architecture and frequently carry out difficult chemical transformations. Manipulation of biosynthetic pathways through in vivo engineering or chemoenzymatic techniques is a promising approach to the generation of novel therapeutics. Presented is our program to elucidate the mechanisms and provide structural information for key steps in the biosynthesis of nonribosomal peptides. The project will examine two important aspects of the biosynthetic methodology: the biosynthesis of nonproteinogenic amino acid building blocks and the selection and loading of amino acids onto the synthetase machinery. We will apply an interdisciplinary combination of X-ray crystallography, organic synthesis and mechanistic enzymology. The results will be applied to the rational engineering of novel amino acid building blocks and is part of our long term goal of understanding the complex mechanisms natural product assembly-line biosynthesis. Systems under study are selected for both exhibited novel enzymology and importance in the biosynthesis of therapeutically important molecules. The proposal describes three enzyme systems, each contributing unique information toward the overall project goals. 1) Nonproteinogenic amino acids are key components of the vancomycin class of antibiotics. An important step in the biosynthesis of 3,5-dihydroxyphenylglycine is the dioxygenation catalyzed by the enzyme DpgC. DpgC is a member of a very small group of cofactor/metal independent oxygenases and has unique chemistry and structure. 2) ?-Amino acids are important building blocks in a wide range of natural and synthetic compounds, including the antitumor/antibiotic enediynes. A novel aminomutase, SgcC4, containing the rare cofactor 4-methylideneimidazolone catalyzes the 1,2-amino shift of ?-tyrosine to generate ?-tyrosine. 3) The structural basis of domain/domain interactions of nonribosomal peptide machinery will be examined using designed synthetic analogs as tethering agents. We will apply this approach to the stand-alone didomain constructs responsible for activation and loading of amino acids.

Public Health Relevance

A large percentage of useful therapeutics are derived from natural sources like microbes and plants. Our research program is interested in understanding the details of how simple organisms developed complex chemistry to produce important drug molecules. This effort will assist in efforts to manipulate natural product producers as a route to the discovery of novel drugs with desired properties.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM086570-06
Application #
8446437
Study Section
Synthetic and Biological Chemistry B Study Section (SBCB)
Program Officer
Gerratana, Barbara
Project Start
2009-04-10
Project End
2015-03-31
Budget Start
2013-04-01
Budget End
2015-03-31
Support Year
6
Fiscal Year
2013
Total Cost
$258,760
Indirect Cost
$82,132
Name
University of Florida
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
969663814
City
Gainesville
State
FL
Country
United States
Zip Code
32611