6L18

XFEL structure of T4dCH D179N mutant complex with natively expressed dTMP

Summary for 6L18

• Entry DOI: 10.2210/pdb6l18/pdb
• Descriptor: Deoxycytidylate 5-hydroxymethyltransferase, THYMIDINE-5'-PHOSPHATE, IODIDE ION, ... (5 entities in total)
• Functional Keywords: xfel, room temperature, dtmp, complex, natively inhibited, hydroxymethylase, transferase
• Biological source: Enterobacteria phage T4
• Total number of polymer chains: 1
• Total formula weight: 30341.78

Authors

Park, S.H.,Song, H.K. (deposition date: 2019-09-27, release date: 2019-12-04, Last modification date: 2023-11-22)

Primary citation

Park, S.H.,Park, J.,Lee, S.J.,Yang, W.S.,Park, S.,Kim, K.,Park, Z.Y.,Song, H.K.
A host dTMP-bound structure of T4 phage dCMP hydroxymethylase mutant using an X-ray free electron laser.
Sci Rep, 9:16316-16316, 2019

PubMed Abstract: The hydroxymethylation of cytosine bases plays a vital role in the phage DNA protection system inside the host Escherichia coli. This modification is known to be catalyzed by the dCMP hydroxymethylase from bacteriophage T4 (T4dCH); structural information on the complexes with the substrate, dCMP and the co-factor, tetrahydrofolate is currently available. However, the detailed mechanism has not been understood clearly owing to a lack of structure in the complex with a reaction intermediate. We have applied the X-ray free electron laser (XFEL) technique to determine a high-resolution structure of a T4dCH D179N active site mutant. The XFEL structure was determined at room temperature and exhibited several unique features in comparison with previously determined structures. Unexpectedly, we observed a bulky electron density at the active site of the mutant that originated from the physiological host (i.e., E. coli). Mass-spectrometric analysis and a cautious interpretation of an electron density map indicated that it was a dTMP molecule. The bound dTMP mimicked the methylene intermediate from dCMP to 5'-hydroxymethy-dCMP, and a critical water molecule for the final hydroxylation was convincingly identified. Therefore, this study provides information that contributes to the understanding of hydroxymethylation.

PubMed: 31705139
DOI: 10.1038/s41598-019-52825-y

Experimental method

X-RAY DIFFRACTION (1.9 Å)