Researchers infer chemical steps involved in methylthiolation of tRNAs

Recent research revealed the chemical steps in a vital cellular modification mechanism that inserts a chemical tag to certain RNAs. Interference of this process in humans can result in cancers, diabetes, and neuronal diseases.

tRNAA new study by Penn State researchers has revealed the chemical steps involved in adding an important tag—a methyl sulfur group—to transfer RNA, a process that, if interfered with in humans, can lead to neuronal diseases, diabetes, and cancers. Image Credit: StudioMolekuul/

A group of scientists headed by chemists from Penn State imaged a protein that expedites this RNA modification in bacteria, enabling scientists to reconstruct the mechanism. The observations of the research elaborating the modification process were published on September 15th, 2021, in the Nature journal.

Transfer RNAs (tRNA) are kinds of RNAs that “read” the genetic code and translate it into a sequence of amino acids to create a protein. The insertion of a chemical tag—a methyl sulfur group—to a specific location on certain tRNAs enhances their capability to translate messenger RNA into proteins.

This modification process is named methylthiolation, and when this does not happen appropriately, errors can be incorporated into the resultant proteins, which in humans causes cancer, neuronal disease, and a heightened risk of developing Type 2 diabetes.

Methylthiolation is ubiquitous across bacteria, plants, and animals. In this study, we determined the structure of a protein called MiaB to better understand its role in facilitating this important modification process in bacteria.”

Squire Booker, Biochemist, The Pennsylvania State University

Squire Booker, who led the research team, is also an investigator with the Howard Hughes Medical Institute.

The MiaB protein from the bacteria Bacteroides uniformis belongs to the radical SAM (S-adenosylmethionine) family of enzymes. Radical SAM enzymes generally employ one of their own iron-sulfur clusters to convert a SAM molecule into a “free radical,” which takes forward the reaction.

Contrary to most other radical SAM enzymes, MiaB consists of two iron-sulfur clusters: a radical SAM cluster and an auxiliary cluster, where the majority of the intricate chemistry occurs.

Imaging MiaB in action with SAM molecules and tRNA at various points at the time of methylthiolation enabled the scientists to deduce the chemical steps taking place during the modification mechanism. Initially, a SAM molecule donates its methyl group to the auxiliary iron-sulfur cluster on MiaB.

The source of the sulfur atom attached to the tRNA has been controversial, but our structures reveal that a methyl group from SAM attaches to a sulfur atom on MiaB’s auxiliary iron-sulfur cluster. This methyl group and the sulfur it attaches to on MiaB are ultimately what transfers to the tRNA, but some additional steps occur before the tRNA can accept the methylthio group.”

Olga Esakova, Study First Author and Assistant Research Professor in Chemistry, The Pennsylvania State University

The insertion of an electron fragments a second molecule of SAM into a free radical. The radical eventually acquires a hydrogen atom from the tRNA, which is replaced with the methylthio group on MiaB.

Initially, the hydrogen on the tRNA is not positioned in a way that allows both access to the radical that removes it and access to the methylthio group that needs to be transferred, because the hydrogen and the atoms attached nearby are all aligned in the same plane.”

Squire Booker, Biochemist, The Pennsylvania State University

Booker also added, “Our structures show that the methylthio group on MiaB’s auxiliary cluster induces a change in geometry at that spot in the tRNA undergoing methylthiolation, which changes into more of a tetrahedral shape, with the hydrogen in an optimal position to be plucked off by the radical and the methylthio group in an optimal position for subsequent transfer.”

The resultant of these steps is tRNA with the inserted methylthio group and an effective modification.

As a further step, the scientists intend to pinpoint how the auxiliary cluster is rebuilt after each turnover so that the mechanism can advance for multiple rounds. They are also analyzing analogous proteins that carry out a similar role in the modification process in humans.

Journal reference:

Esakova, O. A., et al. (2021) Structural basis for tRNA methylthiolation by the radical SAM enzyme MiaB. Nature.


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