8Q5E

Crystal structure of PpSB1-LOV protein from Pseudomonas putida with covalent FMN

Summary for 8Q5E

• Entry DOI: 10.2210/pdb8q5e/pdb
• Descriptor: Sensory box protein, FLAVIN MONONUCLEOTIDE, NICKEL (II) ION (3 entities in total)
• Functional Keywords: covalent flavinylation, signaling protein
• Biological source: Pseudomonas putida KT2440
• Total number of polymer chains: 2
• Total formula weight: 33325.61

Authors

Rozeboom, H.J.,Fraaije, M.W. (deposition date: 2023-08-09, release date: 2023-11-29, Last modification date: 2024-11-06)

Primary citation

Tong, Y.,Kaya, S.G.,Russo, S.,Rozeboom, H.J.,Wijma, H.J.,Fraaije, M.W.
Fixing Flavins: Hijacking a Flavin Transferase for Equipping Flavoproteins with a Covalent Flavin Cofactor.
J.Am.Chem.Soc., 145:27140-27148, 2023

PubMed Abstract: Most flavin-dependent enzymes contain a dissociable flavin cofactor. We present a new approach for installing in vivo a covalent bond between a flavin cofactor and its host protein. By using a flavin transferase and carving a flavinylation motif in target proteins, we demonstrate that "dissociable" flavoproteins can be turned into covalent flavoproteins. Specifically, four different flavin mononucleotide-containing proteins were engineered to undergo covalent flavinylation: a light-oxygen-voltage domain protein, a mini singlet oxygen generator, a nitroreductase, and an old yellow enzyme-type ene reductase. Optimizing the flavinylation motif and expression conditions led to the covalent flavinylation of all four flavoproteins. The engineered covalent flavoproteins retained function and often exhibited improved performance, such as higher thermostability or catalytic performance. The crystal structures of the designed covalent flavoproteins confirmed the designed threonyl-phosphate linkage. The targeted flavoproteins differ in fold and function, indicating that this method of introducing a covalent flavin-protein bond is a powerful new method to create flavoproteins that cannot lose their cofactor, boosting their performance.

PubMed: 38048072
DOI: 10.1021/jacs.3c12009

Experimental method

X-RAY DIFFRACTION (2.4 Å)