9CLH

X-ray diffraction structure of the apo form of papain

Summary for 9CLH

• Entry DOI: 10.2210/pdb9clh/pdb
• Related: 9CKT 9CKW 9CKY 9EG7
• Descriptor: Papain, METHANOL (3 entities in total)
• Functional Keywords: protease, apo, hydrolase
• Biological source: Carica papaya (papaya)
• Total number of polymer chains: 1
• Total formula weight: 23532.34

Authors

Vlahakis, N.W.,Rodriguez, J.A. (deposition date: 2024-07-11, release date: 2025-06-04, Last modification date: 2025-06-11)

Primary citation

Liu, M.,Zang, X.,Vlahakis, N.W.,Rodriguez, J.A.,Ohashi, M.,Tang, Y.
Enzymatic combinatorial synthesis of E-64 and related cysteine protease inhibitors.
Nat.Chem.Biol., 2025

PubMed Abstract: E-64 is an irreversible cysteine protease inhibitor prominently used in chemical biology and drug discovery. Here we uncover a nonribosomal peptide synthetase-independent biosynthetic pathway for E-64, which is widely conserved in fungi. The pathway starts with epoxidation of fumaric acid to the warhead (2S,3S)-trans-epoxysuccinic acid with an Fe(II)/α-ketoglutarate-dependent oxygenase, followed by successive condensation with an L-amino acid by an adenosine triphosphate grasp enzyme and with an amine by the fungal example of amide bond synthetase. Both amide bond-forming enzymes display notable biocatalytic potential, including scalability, stereoselectivity toward the warhead and broader substrate scopes in forming the amide bonds. Biocatalytic cascade with these amide bond-forming enzymes generated a library of cysteine protease inhibitors, leading to more potent cathepsin inhibitors. Additionally, one-pot reactions enabled the preparative synthesis of clinically relevant inhibitors. Our work highlights the importance of biosynthetic investigation for enzyme discovery and the potential of amide bond-forming enzymes in synthesizing small-molecule libraries.

PubMed: 40346252
DOI: 10.1038/s41589-025-01907-2

Experimental method

X-RAY DIFFRACTION (1.6 Å)