Extended Data Fig. 5: Active site interactions of BuMiaB with nucleotides 34 and 35 of the anticodon. | Nature

Extended Data Fig. 5: Active site interactions of BuMiaB with nucleotides 34 and 35 of the anticodon.

From: Structural basis for tRNA methylthiolation by the radical SAM enzyme MiaB

Extended Data Fig. 5

The structure of BuMiaB with the 13-mer RNA and 5’dAH+Met is shown in pink, while the structure of BuMiaB with the 17-mer RNA and SAM is shown in maroon. a, Active site interactions with G34. G34 is often modified, and its base inserts between the TRAM and RS domains in a deep cleft, which provides space for modifications. In the structure of BuMiaB in complex with 5’-dAH+Met and the 13-mer, N10 of G34 is within H-bonding distance to Ser388 of the RS domain. The 2’ and 3’ OH groups are H-bonded to two nitrogen atoms from the guanidinium group of Arg418 from the TRAM-domain. In the structure of BuMiaB with SAM and the 17-mer, the position of G34 is different, and the base no longer interacts with Ser388 and Arg418 (Extended Data Figs. 3b, 4b). b, Active site interactions with A35. In the structure of BuMiaB with 5’-dAH+Met and the 13-mer, the carboxylate oxygens of Asp319 are within H-bonding distance to N6 of A35, while the side-chain of Gln28 is in H-bonding distance to the 2’ OH of A35. The A35 base is π-stacked between Phe348 on one side and the adenine ring of i6A37 on the other. The position of A35 is shifted in the SAM-bound structure and is stabilized by π-stacking with G34 on one side and Phe348 on the other. The rotation of Phe348 supports two different orientations of A35 in the active site of the enzyme. c, Binding of i6A37 in the active site of BuMiaB in the structure with the 13-mer and 5’dAH+Met, showing that the isopentenyl group sits in a hydrophobic patch. All figures have the same colour for the domains and their associated residues: tan for MTTase, grey for radical SAM and green for TRAM.

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