Current Research

Functional Analysis of Enzymes on Biosynthetic Pathways of Plant-derived Bioactive Compounds: 

We use interdisciplinary methods to evaluate enzyme catalysts from various sources, such as bacteria, plants, and yeast, with non-natural substrates. Our vision is to transform natural compounds or synthetically-derived chemicals to novel products. Transfer of the genes encoding these enzymes into a chassis organism can potentially make various bioactive molecules in vivo or in vitro.

Importance of β-Amino Acids as Bioactive Products (more here)

Paclitaxel (Taxol) Pathway Aminomutase. A Taxus phenylalanine aminomutase (TcPAM) converts (2S)-α-phenylalanine ((2S)-α-Phe) to (3R)-β-Phe and lies on the paclitaxel (Taxol™) biosynthetic pathway in Taxus plants.

Andrimid Pathway Aminomutase. TcPAM converts (2S)-α-Phe to (3R)-β-Phe, while PaPAM on the andrimid biosynthetic pathway converts the same substrate to (3S)-β- Phe.

TcPAM must rotate/flip the cinnamate skeleton 180° before exchange and rebinding of the NH2/H pair to the cinnamate, while PaPAM must hold the intermediate stationary (see videos below).

 structure    molecular structure


We used the TcPAM structure in complex with (E)-cinnamate, which functions as both a substrate and an intermediate, and the PaPAM structure to account for the distinct β-amino acid stereochemistries.


Phe455 (spheres) in PaPAM shown displacing the phenylpropanoate ligand (green), preventing a bidentate linkage (magenta) with Arg323. This trajectory may explain the different product stereochemistries of the two enzymes—TcPAM forms a bidentate complex with its substrate.


To further understand how to use, for example, TcPAM and PaPAM chemistry to biocatalyze β-amino acids, it is necessary to understand the subtleties of their mechanisms.

Burst-Phase Analysis. The TcPAM forms a transient MIO-NH2 adduct with a finite lifetime. The lifetime of adduct was unknown for TcPAM or any of the several enzymes in this family until we used stopped-flow monitoring of product release to measure the exponential burst phase at pre-steady state (below).

molecular structure

Hammett correlations. Electron-withdrawing and electron-donating substituents on the aryl ring change the rate-determining step of  the PaPAM-catalyzed reaction. Studies on this family of enzymes (class-I lyase), including PaPAM, began in 1967. Our group showed for the first time that the rate-determining step of a class-1 lyase aminomutase is sensitive to the electronics of the substituents.


The influence of the substituents on the kcat of PaPAM revealed a concavedown or a downward break in correlations with Hammett substituent constants (σ). The trend suggests the rate-determining step changes from the elimination to the amination step based on the electronic properties of the substituent.

Further studies include investigating the substrate selectivity and kinetics of mutant forms of PAM, assessing the cryptic mechanism that yields product stereochemistry, surveying binding isotope effects, and conducting structure/function assignments based on homology and X-ray crystallographic data.

In vivo Biocatalysis of β-Amino Acids



Biocatalysis of Docetaxel (more here)

Taxane analogues (docetaxel, paclitaxel, cabazitaxel, paclitaxel C, and tesetaxel) are used 1) for breast, ovarian, and prostate cancers, 2) to stem complications from stent implants in heart surgery, and 3) to work potentially as neuroprotectants against stroke. Current methods to make docetaxel still use an 11 to 12-step semisynthesis, which involves protecting group chemistry that compromises yields and reduces atom economy.

We use regioselective biocatalysts (Taxus Acyltransferases (AT) and Bacterial CoA Ligases) to bypass protecting group chemistry to make docetaxel.

Streamlined 3-Step Biocatalysis of Docetaxel: An alternative to make docetaxel:

3-step biocatalysis


A new graduate student can embark on studies involving organic chemistry synthesis of novel surrogate substrates. Other areas of training include molecular cloning techniques, expression of various enzymes in E. coli, and assay development. Included are basic biochemical applications and molecular engineering approaches related to enzyme kinetics, enzyme purification and characterization, and various analytical techniques (such as NMR, GC/ MS, LC-MS(/MS), and X-ray crystallography).