Pritam Giri

Konkuk University

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Co-Authors

Hyungdon Yun

Konkuk University

Taresh P. Khobragade

Konkuk University

Sharad Sarak

Konkuk University

Amol D. Pagar

Konkuk University

Hyunwoo Jeon

Konkuk University

Shouvik Chattopadhyay

Jadavpur University

Klaus Harms

Philipps University of Marburg

Mahesh D. Patil

Mahatma Phule Krishi Vidyapeeth

Seonga Lim

Konkuk University

Samim Khan

Aliah University

Cooperative Institutions

Konkuk University

Pohang University of Science and Technology

Seoul National University

Institute for Basic Science

Government of the Republic of Korea

Chong Kun Dang Bio (South Korea)

Konkuk University Medical Center

Korea University

Sungkyunkwan University

National Institute of Pharmaceutical Education and Research

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Creation of a (R)-β-Transaminase by Directed Evolution of d-Amino Acid Aminotransferase

ACS Catalysis (2022)

Hyunwoo Jeon Amol D. Pagar Hyeona Kang Pritam Giri Saravanan Prabhu Nadarajan

β-Transaminases (β-TAs) have shown considerable potential as biocatalysts for the synthesis of chiral β- and γ-amino acid. Herein, a (R)-β-TA was successfully created by the directed evolution of d-amino acid aminotransferase. The specific activities of created (R)-β-TA for β-phenylalanine (1.74 U/mg) and γ-phenylbutanoic acid (1.67 U/mg) were comparable with those of naturally occurring (S)-β-TAs. Moreover, the designed (R)-β-TA also showed potential for the biocatalytic synthesis of a δ-amino acid with specific activity (0.23 U/mg), which has not yet been explored. Enantiopure (S)- and (R)-forms of various β-, γ- and δ-amino acids (>99% ee) were synthesized using engineered (R)-β-TA-mediated kinetic resolution and asymmetric synthesis, respectively.

Enantiopure drug

Transaminase

Kinetic resolution

10.1021/acscatal.2c04221

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Citations (8)

Biocatalytic Cascade for Synthesis of Sitagliptin Intermediate Employing Coupled Transaminase

Biotechnology and Bioprocess Engineering (2023)

Taresh P. Khobragade Amol D. Pagar Pritam Giri Sharad Sarak Hyunwoo Jeon

Enantiopure drug

Biotransformation

Esterase

Biocatalysis

Acetophenone

Transaminase

Cascade reaction

10.1007/s12257-023-0085-5

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Citations (5)

Exemplifying Natural (R)-β-Transamination Potential of Fold Type-IV Transaminase for Kinetic Resolution of rac-β-Amino Acids Synthesized from Aldehydes

ACS Sustainable Chemistry & Engineering (2024)

Amol D. Pagar Taresh P. Khobragade Pritam Giri Hyunwoo Jeon Sungchan Cho

Sustainable synthesis of chiral β-amino acids from renewable feedstocks is of significant importance. The Rodionov reaction is a popular method to synthesize rac-β-amino acids from aldehydes, which are abundant byproducts of biomass. rac-β-Amino acids are easily converted into the enantiopure (R)-form by kinetic resolution using (S)-β-transaminases (TAs). However, the inaccessibility of natural (R)-β-transaminases with a broader substrate scope limits the application of the Rodionov reaction to produce (S)-β-amino acids. Here, we report the kinetic resolution of rac-β-amino acids by employing fold type-IV transaminase from Thermobaculum terrenum (TATT) with natural (R)-β-transaminase activity. Various rac-β-amino acids were successfully resolved by TATT into (S)-form with excellent conversions (∼50%) and enantiomeric excess (>99%) using pyruvate as an amino acceptor. The directed evolution of TATT resulted in a variant M-58 (K112F/F114M) with ∼2-fold higher activity and was able to perform large-scale kinetic resolution of 20 mM rac-3-amino-3-(3-fluorophenyl) propanoic acid (1h) with complete conversion and enantiomeric excess >99% within 24 h.

Transamination

Kinetic resolution

Transaminase

10.1021/acssuschemeng.4c00794

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Non-Canonical Amino Acid-Based Engineering of (R)-Amine Transaminase

Frontiers in Chemistry (2022)

Amol D. Pagar Hyunwoo Jeon Taresh P. Khobragade Sharad Sarak Pritam Giri

Non-canonical amino acids (ncAAs) have been utilized as an invaluable tool for modulating the active site of the enzymes, probing the complex enzyme mechanisms, improving catalytic activity, and designing new to nature enzymes. Here, we report site-specific incorporation of p -benzoyl phenylalanine ( p BpA) to engineer ( R )-amine transaminase previously created from d -amino acid aminotransferase scaffold. Replacement of the single Phe88 residue at the active site with p BpA exhibits a significant 15-fold and 8-fold enhancement in activity for 1-phenylpropan-1-amine and benzaldehyde, respectively. Reshaping of the enzyme’s active site afforded an another variant F86A/F88 p BpA, with 30% higher thermostability at 55°C without affecting parent enzyme activity. Moreover, various racemic amines were successfully resolved by transaminase variants into ( S )-amines with excellent conversions (∼50%) and enantiomeric excess (>99%) using pyruvate as an amino acceptor. Additionally, kinetic resolution of the 1-phenylpropan-1-amine was performed using benzaldehyde as an amino acceptor, which is cheaper than pyruvate. Our results highlight the utility of ncAAs for designing enzymes with enhanced functionality beyond the limit of 20 canonical amino acids.

Transaminase

Kinetic resolution

Thermostability

Peptidomimetic

10.3389/fchem.2022.839636

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Citations (17)

Dual-function transaminases with hybrid nanoflower for the production of value-added chemicals from biobased levulinic acid

Frontiers in Bioengineering and Biotechnology (2023)

Taresh P. Khobragade Pritam Giri Amol D. Pagar Mahesh D. Patil Sharad Sarak

The U.S. Department of Energy has listed levulinic acid (LA) as one of the top 12 compounds derived from biomass. LA has gained much attention owing to its conversion into enantiopure 4-aminopentanoic acid through an amination reaction. Herein, we developed a coupled-enzyme recyclable cascade employing two transaminases (TAs) for the synthesis of ( S )-4-aminopentanoic acid. TAs were first utilized to convert LA into ( S )-4-aminopentanoic acid using ( S )-α-Methylbenzylamine [( S )-α-MBA] as an amino donor. The deaminated ( S )-α-MBA i.e., acetophenone was recycled back using a second TAs while using isopropyl amine (IPA) amino donor to generate easily removable acetone. Enzymatic reactions were carried out using different systems, with conversions ranging from 30% to 80%. Furthermore, the hybrid nanoflowers (HNF) of the fusion protein were constructed which afforded complete biocatalytic conversion of LA to the desired ( S )-4-aminopentanoic acid. The created HNF demonstrated storage stability for over a month and can be reused for up to 7 sequential cycles. A preparative scale reaction (100 mL) achieved the complete conversion with an isolated yield of 62%. Furthermore, the applicability of this recycling system was tested with different β-keto ester substrates, wherein 18%–48% of corresponding β-amino acids were synthesized. Finally, this recycling system was applied for the biosynthesis of pharmaceutical important drug sitagliptin intermediate (( R )-3-amino-4-(2,4,5-triflurophenyl) butanoic acid) with an excellent conversion 82%.

Enantiopure drug

Levulinic acid

Isopropyl alcohol

Reductive amination

10.3389/fbioe.2023.1280464

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Observation of ultrafast electrons in pendant-embedded conducting two-dimensional polymers

Chem (2024)

Yeonsang Lee Minhyuk Choi Ina Park In‐Chul Hwang Sk. Atiur Rahaman

Electron Mobility

10.1016/j.chempr.2023.12.007

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Citations (1)

Promoter engineering‐mediated Tuning of esterase and transaminase expression for the chemoenzymatic synthesis of sitagliptin phosphate at the kilogram‐scale

Biotechnology and Bioengineering (2021)

Taresh P. Khobragade Seongseon Yu Hyunsang Jung Mahesh D. Patil Sharad Sarak

Here, we report a bienzymatic cascade to produce β-amino acids as an intermediate for the synthesis of the leading oral antidiabetic drug, sitagliptin. A whole-cell biotransformation using recombinant Escherichia coli coexpressing a esterase and transaminase were developed, wherein the desired expression level of each enzyme was achieved by promotor engineering. The small-scale reactions (30 ml) performed under optimized conditions at varying amounts of substrate (100-300 mM) resulted in excellent conversions of 82%-95% for the desired product. Finally, a kilogram-scale enzymatic reaction (250 mM substrate, 220 L) was carried out to produce β-amino acid (229 mM). Sitagliptin phosphate was chemically synthesized from β-amino acids with 82% yield and > 99% purity.

Transaminase

Biotransformation

Esterase

Biocatalysis

10.1002/bit.27819

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Citations (15)

CCDC 1940162: Experimental Crystal Structure Determination

The Cambridge Structural Database (2019)

Tamal Dutta Saikat Mirdya Pritam Giri Shouvik Chattopadhyay

10.5517/ccdc.csd.cc233ww0

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Biocatalysis enables the scalable conversion of biobased furans into various furfurylamines

Nature Communications (2024)

Pritam Giri Seonga Lim Taresh P. Khobragade Amol D. Pagar Mahesh D. Patil

Biobased furans have emerged as chemical building blocks for the development of materials because of their diverse scaffolds and as they can be directly prepared from sugars. However, selective, efficient, and cost-effective scalable conversion of biobased furans remains elusive. Here, we report a robust transaminase (TA) from Shimia marina (SMTA) that enables the scalable amination of biobased furanaldehydes with high activity and broad substrate specificity. Crystallographic and mutagenesis analyses provide mechanistic insights and a structural basis for understanding SMTA, which enables a higher substrate conversion. The enzymatic cascade process established in this study allows one-pot synthesis of 2,5-bis(aminomethyl)furan (BAMF) and 5-(aminomethyl)furan-2-carboxylic acid from 5-hydroxymethylfurfural. The biosynthesis of various furfurylamines, including a one-pot cascade reaction for BAMF generation using whole cells, demonstrates their practical application in the pharmaceutical and polymer industries.

Biocatalysis

10.1038/s41467-024-50637-x

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Citations (2)

Synthesis, characterization and DFT study on two copper(II) complexes with a naphthalene-based Schiff base: Examples of stronger chelate–chelate interactions than those reported for classical π–π complexes

Polyhedron (2018)

Samim Khan Pritam Giri Antonio Bauzá Klaus Harms Antonio Frontera

Dicyanamide

Thiocyanate

10.1016/j.poly.2018.09.074

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Citations (11)