6OZ1

Crystal structure of the adenylation (A) domain of the carboxylate reductase (CAR) GR01_22995 from Mycobacterium chelonae

Summary for 6OZ1

• Entry DOI: 10.2210/pdb6oz1/pdb
• Descriptor: Oxidoreductase, SULFATE ION, CHLORIDE ION, ... (7 entities in total)
• Functional Keywords: adenylation domain, carboxylate reductase, oxidoreductase
• Biological source: Mycobacterium chelonae
• Total number of polymer chains: 1
• Total formula weight: 72024.14

Authors

Stogios, P.J.,Evdokimova, E.,Di Leo, R.,Fedorchuk, T.,Khusnutdinova, A.,Yakunin, A.F.,Savchenko, A. (deposition date: 2019-05-15, release date: 2020-04-22, Last modification date: 2023-10-11)

Primary citation

Fedorchuk, T.P.,Khusnutdinova, A.N.,Evdokimova, E.,Flick, R.,Di Leo, R.,Stogios, P.,Savchenko, A.,Yakunin, A.F.
One-Pot Biocatalytic Transformation of Adipic Acid to 6-Aminocaproic Acid and 1,6-Hexamethylenediamine Using Carboxylic Acid Reductases and Transaminases.
J.Am.Chem.Soc., 142:1038-1048, 2020

PubMed Abstract: Production of platform chemicals from renewable feedstocks is becoming increasingly important due to concerns on environmental contamination, climate change, and depletion of fossil fuels. Adipic acid (AA), 6-aminocaproic acid (6-ACA) and 1,6-hexamethylenediamine (HMD) are key precursors for nylon synthesis, which are currently produced primarily from petroleum-based feedstocks. In recent years, the biosynthesis of adipic acid from renewable feedstocks has been demonstrated using both bacterial and yeast cells. Here we report the biocatalytic conversion/transformation of AA to 6-ACA and HMD by carboxylic acid reductases (CARs) and transaminases (TAs), which involves two rounds (cascades) of reduction/amination reactions (AA → 6-ACA → HMD). Using purified wild type CARs and TAs supplemented with cofactor regenerating systems for ATP, NADPH, and amine donor, we established a one-pot enzyme cascade catalyzing up to 95% conversion of AA to 6-ACA. To increase the cascade activity for the transformation of 6-ACA to HMD, we determined the crystal structure of the CAR substrate-binding domain in complex with AMP and succinate and engineered three mutant CARs with enhanced activity against 6-ACA. In combination with TAs, the CAR L342E protein showed 50-75% conversion of 6-ACA to HMD. For the transformation of AA to HMD (via 6-ACA), the wild type CAR was combined with the L342E variant and two different TAs resulting in up to 30% conversion to HMD and 70% to 6-ACA. Our results highlight the suitability of CARs and TAs for several rounds of reduction/amination reactions in one-pot cascade systems and their potential for the biobased synthesis of terminal amines.

PubMed: 31886667
DOI: 10.1021/jacs.9b11761

Experimental method

X-RAY DIFFRACTION (1.97 Å)