Biology subjects

Chang, M. C. Y.

Publications and source records attributed to Chang, M. C. Y..

4 recordsLinked to original sources

bioRxiv2023

Biosynthesis of Strained Amino Acids Through a PLP-Dependent Enzyme via Cryptic Halogenation

Amino acids (AAs) are modular and modifiable building blocks which nature uses to synthesize both macromolecules, such as proteins, and small molecule natural products, such as alkaloids and non-ribosomal peptides (NRPs). While the 20 main proteinogenic AAs display relatively limited side-chain diversity, a wide range of non-canonical amino acids (ncAAs) exist that are not used by the ribosome for protein synthesis but contain a broad array of structural features and functional groups not found in proteinogenic AAs. In this communication, we report the discovery of the biosynthetic pathway for a new ncAA, pazamine, which contains a cyclopropane ring formed in two steps. In the first step, a chlorine is added onto the C4 position of lysine by a radical halogenase PazA. The cyclopropane ring is then formed in the next step by a pyridoxal-5-phosphate-dependent enzyme, PazB, via an SN2-like attack onto C4 to eliminate chloride. Genetic studies of this pathway in the native host, Pseudomonas azotoformans, show that pazamine and its succinylated derivative, pazamide, potentially inhibit ethylene biosynthesis in growing plants based on alterations in the root phenotype of Arabidopsis thaliana seedlings. We further show that PazB can be utilized to make an alternative cyclobutane-containing AA. These discoveries may lead to advances in biocatalytic production of specialty chemicals and agricultural biotechnology.

biochemistry↗

bioRxiv2023

A Cellular Platform for Production of C4 Monomers

Living organisms carry out a wide range of remarkable functions, including the synthesis of thousands of simple and complex chemical structures for cellular growth and maintenance. The manipulation of this reaction network has allowed for the genetic engineering of cells for targeted chemical synthesis, but it remains challenging to alter the program underlying their fundamental chemical behavior. By taking advantage of the unique ability of living systems to use evolution to find solutions to complex problems, we have achieved ~95% theoretical yield of three C4 commodity chemicals, n-butanol, 1,3-butanediol, and 4-hydroxy-2-butanone. Genomic sequencing of the evolved strains identified pcnB and rpoBC as two gene loci that are able to alter carbon flow by remodeling the transcriptional landscape of the cell, highlighting the potential of synthetic pathways as a tool to identify metabolic control points.

synthetic biology↗

bioRxiv2022

Regioselective control of biocatalytic C-H activation and halogenation

Biocatalytic C-H activation has the potential to merge enzymatic and synthetic strategies for bond formation. FeII/KG-dependent halogenases are particularly distinguished for their ability both to control selective C-H activation as well as to direct group transfer of a bound anion along a reaction axis separate from oxygen rebound, enabling the development of new transformations. In this context, we elucidate the basis for selectivity of enzymes that perform selective halogenation to yield 4-Cl-lysine (BesD), 5-Cl-lysine (HalB), and 4-Cl-ornithine (HalD), allowing us to probe how regioselectivity and chain length selectivity are achieved. We now report the crystal structure of the HalB and HalD, revealing the key role of the substrate-lid in positioning the substrate for C4 vs C5 chlorination and recognition of lysine vs ornithine. Targeted engineering of the substrate-binding lid further demonstrates that these selectivities can be altered or switched, showcasing the potential to develop halogenases for biocatalytic applications.

biochemistry↗

bioRxiv2020

A Hybrid Cellular and Heterogeneous Catalyst Strategy for the Production of Olefins from Glucose

Living systems provide a promising approach to chemical synthesis, having been optimized by evolution to convert renewable carbon sources such as glucose to an enormous range of small molecules. However, a large number of synthetic structures can still be difficult to obtain solely from cells, such as unsubstituted hydrocarbons. In this work, we demonstrate the use of a hybrid cellular-heterogeneous catalytic strategy to produce olefins from glucose, using a selective hydrolase to generate an activated intermediate that is readily deoxygenated. Using a new family of iterative thiolase enzymes, we have genetically engineered a microbial strain that produces 4.3 {+/-} 0.4 g L-1 of fatty acid from glucose with 86% captured as 3-hydroxyoctanoic and 3-hydroxydecanoic acids. This 3-hydroxy substituent serves as a leaving group enabling heterogeneous tandem decarboxylation-dehydration routes to olefinic products on Lewis acidic catalysts without the additional redox input required for enzymatic or chemical deoxygenation of simple fatty acids.

synthetic biology↗