Structural Basis for Methyl Transfer by a Radical SAM Enzyme
Amy Rosenzweig group (Northwestern University) and collaborators
Antibiotic drug resistance is a growing global public health
problem. In the bacterium Staphylococcus aureus, the enzyme Cfr
confers resistance to antibiotics by methylating the C8 position of adenosine
2503 (A2503), which is ultimately found in the peptidyl transferase center of
the 50S subunit of the bacterial ribosome. Cfr is closely related to RlmN,
which modifies the C2 position on A2503 for a housekeeping function unrelated
to drug resistance. These two enzymes use Sadenosylmethionine (SAM) as both
the source of the methyl group and the source of a 5'-deoxyadenosyl
5'-radical (5'-dA·) generated using a [4Fe-4S] cluster. In the
proposed mechanism, the enzyme first appends a methyl group to a conserved
cysteine in the active site (Cys 355 in RlmN) in a standard SAM-dependent
methyl transfer reaction. A second molecule of SAM is then reductively
cleaved to activate the Cys-appended methyl group which is transferred to the
appropriate carbon atom on A2503. Crystal structures of RlmN determined by
researchers in the labs of Amy Rosenzweig and Squire Booker (Pennsylvania
State University) provide molecular-level insight into this unusual
mechanism. In the 2.05-Å resolution structure of Escherichia
coli RlmN with bound SAM, the SAM cosubstrate is coordinated to a
[4Fe-4S] cluster located within an active site architecture typical of
radical SAM superfamily enzymes. The structure confirms the presence of a
methyl substituent on Cys355, well poised for a subsequent hydrogen atom
abstraction reaction to activate the methyl group. The active site
arrangement also suggests an identical SAM binding site for the initial
methyl transfer to Cys355, a reaction not usually catalyzed by SAM bound to
an iron-sulfur cluster. Cys355 lies within a flexible loop that dips into
the active site, providing a structural rationale for its multifunctional
role in the reaction. A large positively charged opening to the active site
provides an obvious approach for a bulky ribosomal RNA substrate. Finally,
the RlmN structures help in understanding how Cfr might have evolved to
facilitate antibiotic resistance.
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Figure: The RlmN active site. The
[4Fe-4S] cluster is shown as orange and yellow spheres and the SAM
cosubstrate is shown as sticks. Methylated residue Cys 355, shown as sticks,
derives from a flexible loop region shown in white. The rest of the protein
is shown as a blue surface. |
Citation: Boal AK, Grove TL, McLaughlin MI, Yennawar NH,
Booker SJ, Rosenzweig AC. Structural basis for methyl transfer by a
radical SAM enzyme. Science 2011 May 27; 332: 1089-92. doi: 10.1126/science.1205358.
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