Joo-Hyun Seo

Bio: Joo-Hyun Seo is an academic researcher from Sun Moon University. The author has contributed to research in topics: Transaminase & Biotransformation. The author has an hindex of 19, co-authored 45 publications receiving 1080 citations. Previous affiliations of Joo-Hyun Seo include Kookmin University & Korea Institute for Advanced Study.

Redesigning the substrate specificity of ω‐aminotransferase for the kinetic resolution of aliphatic chiral amines

Abstract: Substrate specificity of the omega-aminotransferase obtained from Vibrio fluvialis (omega-ATVf) was rationally redesigned for the kinetic resolution of aliphatic chiral amines. omega-ATVf showed unique substrate specificity toward aromatic amines with a high enantioselectivity (E > 100) for (S)-enantiomers. However, the substrate specificity of this enzyme was much narrower toward aliphatic amines. To overcome the narrow substrate specificity toward aliphatic amines, we redesigned the substrate specificity of omega-ATVf using homology modeling and the substrate structure- activity relationship. The homology model and the substrate structure-activity relationship showed that the active site of omega-ATVf consists of one large substrate-binding site and another small substrate-binding site. The key determinant in the small substrate-binding site was D25, whose role was expected to mask R415 and to generate the electrostatic repulsion with the substrate's alpha-carboxylate group. In the large substrate-binding site, R256 was predicted to recognize the alpha-carboxylate group of substrate thus obeying the dual substrate recognition mechanism of aminotransferase subgroup II enzymes. Among the several amino acid residues in the large substrate-binding site, W57 and W147, with their bulky side chains, were expected to restrict the recognition of aliphatic amines. Two mutant enzymes, W57G and W147G, showed significant changes in their substrate specificity such that they catalyzed transamination of a broad range of aliphatic amines without losing the original activities toward aromatic amines and enantioselectivity.

Cloning and Characterization of a Novel β-Transaminase from Mesorhizobium sp. Strain LUK: a New Biocatalyst for the Synthesis of Enantiomerically Pure β-Amino Acids

Abstract: β-Amino acids occur naturally in diverse forms. They have been found as free forms of metabolites, such as β-alanine and β-aminobutyric acid, in mammals and lower organisms (19). They are also found as a component of peptidic or nonpeptidic molecules (28). β-Amino acid-containing molecules have interesting pharmacological roles such as antibiotics (51), antitumor agents (14, 36, 53), or antifungal agents (15). β-Lactams and taxoids are such examples of the important molecules further developed from the natural compounds with β-amino acid moiety (26, 38). The higher stability of β-amino acid peptides against peptidases also has drawn great interest in peptide chemistry (16). The significance of β-amino acids and their related molecules attracts increasing attention for the synthesis of β-amino acids (5, 37, 38, 41), especially for the enantioselective synthesis of β-amino acids (26, 28, 30, 40). Although recent reviews of the biocatalytic synthesis of β-amino acids elaborated a wide range of approaches, most of the methods dealt with the chiral resolution of precursors using hydrolytic enzymes (29). As an alternative, we have focused on the study of transaminases for the synthesis of versatile chiral amine compounds including unnatural amino acids. Transaminases have been studied for the production of chiral amino acids because they generally show rapid reaction rates, broad substrate specificity, and no requirement for external cofactor regeneration (44, 49, 54). Moreover, transaminases allow asymmetric synthesis from prochiral ketone compounds depending upon the properties of target chemical compounds (1, 4, 10, 11, 17, 49, 50). Though transaminases are not widespread, we have reported an ω-transaminase of Alcaligenes denitrificans which can catalyze mainly the transamination between aliphatic β-amino acids and pyruvate (56). Other examples were recently introduced for the transamination of aliphatic and aromatic β-amino acids by Alcaligenes eutrophus and Variovorax paradoxus (6). Recently, we reported screening of a transaminase having activity toward a β-amino acid and its N-terminal amino acid sequence (27). We have tried to clone the gene of this enzyme by PCR using the degenerative primers of consensus transaminase sequences (56). However, due to the large population of the homologous transaminases in the screened organism, we found a few transaminases without any activities for β-amino acids, even though we used genuine N-terminal amino acid sequence to make one of the degenerative primers. Due to the low recovery of the purified protein, our attempts to obtain any internal peptide sequences had been also unsuccessful. The present study illustrates the easy isolation of enzyme with higher recovery, molecular cloning, sequencing, heterologous expression of the gene encoding the new β-transaminase in Escherichia coli, and its characterization. In this article, we report a novel β-transaminase from Mesorhizobium sp. strain LUK showing a high activity for β-aminocarboxylic acids. In addition, the asymmetric synthesis of a β-aminocarboxylic acid is presented along with a short discussion on the substrate recognition mode of β-transaminase.

High accuracy template based modeling by global optimization

Abstract: For high-accuracy template-based-modelingof CASP7 targets, we have applied a proce-dure based on the rigorous optimization ofscore functions at three stages: multiplealignment, chain building, and side-chainmodeling. We applied the conformationalspace annealing method to a newly developedconsistency based score function for multiplealignment. For chain building, we optimizedthe MODELLER energy function. For side-chain modeling, we optimized a SCWRL-likeenergy function using a rotamer libraryconstructed specifically for a given targetsequence. By rigorous optimization, we haveachieved significant improvement in back-bone as well as side-chain modeling for TBMand TBM/HA targets. For most TBM/HA tar-gets (17/26), the predicted model was moreaccurate than the model one can constructfrom the best template in a posteriori fash-ion. It appears that the current method canextract relevant information out of multipletemplates. Proteins 2007; 69(Suppl 8):83–89. VV C 2007 Wiley-Liss, Inc.

Asymmetric synthesis of L‐homophenylalanine by equilibrium‐shift using recombinant aromatic L‐amino acid transaminase

Abstract: L-Homophenylalanine (L-HPA) was asymmetrically synthesized from 2-oxo-4-phenylbutyric acid (2-OPBA) and L-aspartate using a recombinant aromatic amino acid transaminase (AroAT). To screen microorganisms having such an L-specific AroAT with a relaxed substrate inhibition in the asymmetric synthesis of unnatural amino acids, enrichment cultures were performed in a minimal media containing 50 mM L-HPA as a sole nitrogen source. To reduce the intracellular background synthetic activity by amino acid pools in the cells, a two-step screening method was used. The putative AroAT (i.e., AroATEs) from the screened Enterobacter sp. BK2K-1 was cloned, sequenced, and overexpressed in E. coli cells. The activity of the overexpressed AroATEs was 314-fold higher than that of the wild-type cell. The substrate specificities of the enzyme and homology search revealed that the cloned transaminase is true AroAT. The AroATEs showed a substrate inhibition by 2-OPBA from 40 mM in the asymmetric synthesis, which made it difficult to perform batch asymmetric synthesis of L-HPA at high concentrations of 2-OPBA. To avoid the substrate inhibition by 2-OPBA, intermittent addition of the solid-state substrate was attempted to obtain a high concentration of L-HPA. By using the cell extract (75 U) obtained from the recombinant E. coli harboring the AroATEs gene, the asymmetric synthesis of L-HPA at 840 mM of 2-OPBA resulted in >94% of conversion yield and >99% ee of L-HPA of optical purity. Due to the low solubility ( 99% ee) was easily recovered by simple pH shift of the reaction media. This method can permit very efficient asymmetric synthesis of other unnatural amino acids using a single transaminase reaction.

Automated comparative protein structure modeling with SWISS-MODEL and Swiss-PdbViewer: a historical perspective.

Abstract: SWISS-MODEL pioneered the field of automated modeling as the first protein modeling service on the Internet. In combination with the visualization tool Swiss-PdbViewer, the Internet-based Workspace and the SWISS-MODEL Repository, it provides a fully integrated sequence to structure analysis and modeling platform. This computational environment is made freely available to the scientific community with the aim to hide the computational complexity of structural bioinformatics and encourage bench scientists to make use of the ever-increasing structural information available. Indeed, over the last decade, the availability of structural information has significantly increased for many organisms as a direct consequence of the complementary nature of comparative protein modeling and experimental structure determination. This has a very positive and enabling impact on many different applications in biomedical research as described in this paper.

Biocatalytic Asymmetric Synthesis of Chiral Amines from Ketones Applied to Sitagliptin Manufacture

Abstract: Pharmaceutical synthesis can benefit greatly from the selectivity gains associated with enzymatic catalysis Here, we report an efficient biocatalytic process to replace a recently implemented rhodium-catalyzed asymmetric enamine hydrogenation for the large-scale manufacture of the antidiabetic compound sitagliptin Starting from an enzyme that had the catalytic machinery to perform the desired chemistry but lacked any activity toward the prositagliptin ketone, we applied a substrate walking, modeling, and mutation approach to create a transaminase with marginal activity for the synthesis of the chiral amine; this variant was then further engineered via directed evolution for practical application in a manufacturing setting The resultant biocatalysts showed broad applicability toward the synthesis of chiral amines that previously were accessible only via resolution This work underscores the maturation of biocatalysis to enable efficient, economical, and environmentally benign processes for the manufacture of pharmaceuticals

Improving physical realism, stereochemistry, and side-chain accuracy in homology modeling: Four approaches that performed well in CASP8.

Abstract: A correct alignment is an essential requirement in homology modeling. Yet in order to bridge the structural gap between template and target, which may not only involve loop rearrangements, but also shifts of secondary structure elements and repacking of core residues, high-resolution refinement methods with full atomic details are needed. Here, we describe four approaches that address this "last mile of the protein folding problem" and have performed well during CASP8, yielding physically realistic models: YASARA, which runs molecular dynamics simulations of models in explicit solvent, using a new partly knowledge-based all atom force field derived from Amber, whose parameters have been optimized to minimize the damage done to protein crystal structures. The LEE-SERVER, which makes extensive use of conformational space annealing to create alignments, to help Modeller build physically realistic models while satisfying input restraints from templates and CHARMM stereochemistry, and to remodel the side-chains. ROSETTA, whose high resolution refinement protocol combines a physically realistic all atom force field with Monte Carlo minimization to allow the large conformational space to be sampled quickly. And finally UNDERTAKER, which creates a pool of candidate models from various templates and then optimizes them with an adaptive genetic algorithm, using a primarily empirical cost function that does not include bond angle, bond length, or other physics-like terms.

A comprehensive review on tyrosinase inhibitors.

Abstract: Tyrosinase is a multi-copper enzyme which is widely distributed in different organisms and plays an important role in the melanogenesis and enzymatic browning. Therefore, its inhibitors can be attractive in cosmetics and medicinal industries as depigmentation agents and also in food and agriculture industries as antibrowning compounds. For this purpose, many natural, semi-synthetic and synthetic inhibitors have been developed by different screening methods to date. This review has focused on the tyrosinase inhibitors discovered from all sources and biochemically characterised in the last four decades.