7EPN

Ketosteroid Isomerase KSI native

Summary for 7EPN

• Entry DOI: 10.2210/pdb7epn/pdb
• Descriptor: SnoaL-like domain-containing protein (2 entities in total)
• Functional Keywords: kemp elimination, msksi, mycobacterium smegmatis, crystal structure., isomerase
• Biological source: Mycolicibacterium smegmatis (strain ATCC 700084 / mc(2)155) (Mycobacterium smegmatis)
• Total number of polymer chains: 2
• Total formula weight: 32788.66

Authors

Liang, Y.,Zhang, Q.,Bartlam, M. (deposition date: 2021-04-27, release date: 2021-12-22, Last modification date: 2023-11-29)

Primary citation

Liang, Y.,Li, W.,Liang, H.,Lou, X.,Liu, R.,Zhang, Q.,Bartlam, M.
Structural characterization and Kemp eliminase activity of the Mycobacterium smegmatis Ketosteroid Isomerase.
Biochem.Biophys.Res.Commun., 560:159-164, 2021

PubMed Abstract: The Kemp elimination reaction, involving the ring-opening of benzoxazole and its derivatives under the action of natural enzymes or chemical catalysts, has been of interest to researchers since its discovery. Because this reaction does not exist in all currently known metabolic pathways, the computational design of Kemp eliminases has provided valuable insights into principles of enzymatic catalysis. However, it was discovered that the naturally occurring promiscuous enzymes ydbC, xapA and ketosteroid isomerase also can catalyze Kemp elimination. Here, we report the crystal structure of ketosteroid isomerase (KSI) from Mycobacterium smegmatis MC2 155. MsKSI crystallizes in the P222 space group with two molecules in an asymmetric unit, and ultracentrifugation data confirms that it forms a stable dimer in solution, consistent with the 1.9 Å-resolution structure. Our assays confirm that MsKSI accelerates the Kemp elimination of 5-nitrobenzoxazole (5NBI) with an optimal pH of 5.5. A 2.35 Å resolution crystal structure of the MsKSI-5NBI complex reveals that the substrate 5NBI is bound in the active pocket of the enzyme composed of hydrophobic residues. In addition, the Glu127 residue is proposed to play an important role as a general base in proton transfer and breaking weak O-N bonds to open the five-membered ring. This work provides a starting point for exploring the artificial modification of MsKSI using the natural enzyme as the backbone.

PubMed: 33992958
DOI: 10.1016/j.bbrc.2021.05.007

Experimental method

X-RAY DIFFRACTION (1.9 Å)