title: Radical-mediated enzymatic methylation: a tale of two SAMS.
abstract: Methylation is an essential and ubiquitous reaction that plays an important role in a wide range of biological processes. Most biological methylations use S-adenosylmethionine (SAM) as the methyl donor and proceed via an S(N)2 displacement mechanism. However, researchers have discovered an increasing number of methylations that involve radical chemistry. The enzymes known to catalyze these reactions all belong to the radical SAM superfamily. This family of enzymes utilizes a specialized [4Fe-4S] cluster for reductive cleavage of SAM to yield a highly reactive 5'-deoxyadenosyl (dAdo) radical. Radical chemistry is then imposed on a variety of organic substrates, leading to a diverse array of transformations. Until recently, researchers had not fully understood how these enzymes employ radical chemistry to mediate a methyl transfer reaction.  Sequence analyses reveal that the currently identified radical SAM methyltransferases (RSMTs) can be grouped into three classes, which appear distinct in protein architecture and mechanism. Class A RSMTs mainly include the rRNA methyltransferases RlmN and Cfr from various origins. As exemplified by  Escherichia coli  RlmN, these proteins have a single canonical radical SAM core domain that includes an (βα)(6) partial barrel most similar to that of pyruvate formate lyase-activase. The exciting recent studies on RlmN and Cfr are beginning to provide insights into the intriguing chemistry of class A RSMTs. These enzymes utilize a methylene radical generated on a unique methylated cysteine residue. However, based on the variety of substrates used by the other classes of RSMTs, alternative mechanisms are likely to be discovered. Class B RSMTs contain a proposed N-terminal cobalamin binding domain in addition to a radical SAM domain at the C-terminus. This class of proteins methylates diverse substrates at inert sp(3) carbons, aromatic heterocycles, and phosphinates, possibly involving a cobalamin-mediated methyl transfer process. Class C RSMTs share significant sequence similarity with coproporphyrinogen III oxidase HemN. Despite methylating similar substrates (aromatic heterocycles), class C RSMTs likely employ a mechanism distinct from that of class A because two conserved cysteines that are required for class A are typically not found in class C RSMTs.  Class A and class B enzymes probably share the use of two molecules of SAM: one to generate a dAdo radical and one to provide the methyl group to the substrate. In class A, a cysteine would act as a conduit of the methyl group whereas in class B cobalamin may serve this purpose. Currently no clues are available regarding the mechanism of class C RSMTs, but the sequence similarities between its members and HemN and the observation that HemN binds two SAM molecules suggest that class C enzymes could use two SAM molecules for catalysis. The diverse strategies for using two SAM molecules reflect the rich chemistry of radical-mediated methylation reactions and the remarkable versatility of the radical SAM superfamily.
authors.affiliation: Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, Illinois 61801, USA.
authors.forename: Qi
authors.initials: Q
authors.lastname: Zhang
authors.forename: Wilfred A
authors.initials: WA
authors.lastname: van der Donk
authors.forename: Wen
authors.initials: W
authors.lastname: Liu
mesh_terms: D005609:Free Radicals / Q000737:chemistry; D008745:Methylation; D008780:Methyltransferases / Q000737:chemistry / Q000378:metabolism*; D012436:S-Adenosylmethionine / Q000737:chemistry* / Q000378:metabolism; D014805:Vitamin B 12 / Q000378:metabolism
chemical_list: D005609:Free Radicals; D012436:S-Adenosylmethionine; D008780:Methyltransferases; D014805:Vitamin B 12
references.citation: J Biol Chem. 2004 Nov 12;279(46):47555-63
references.pmid: 15339930
references.citation: J Biol Chem. 2011 Jun 17;286(24):21287-94
references.pmid: 21454624
references.citation: Chem Rev. 2011 Apr 13;111(4):2487-506
references.pmid: 21370834
references.citation: J Am Chem Soc. 2009 Jul 8;131(26):9347-53
references.pmid: 19514719
references.citation: J Am Chem Soc. 2009 Apr 1;131(12):4327-34
references.pmid: 19265401
references.citation: Science. 2011 May 27;332(6033):1089-92
references.pmid: 21527678
references.citation: Chem Biol. 2009 Feb 27;16(2):141-7
references.pmid: 19246004
references.citation: J Antibiot (Tokyo). 2004 Jul;57(7):436-45
references.pmid: 15376556
references.citation: Antimicrob Agents Chemother. 2005 Jan;49(1):230-40
references.pmid: 15616300
references.citation: J Biol Chem. 1980 Oct 10;255(19):9124-7
references.pmid: 6997310
references.citation: Nat Chem Biol. 2011 Mar;7(3):154-60
references.pmid: 21240261
references.citation: Acc Chem Res. 2001 Aug;34(8):681-9
references.pmid: 11513576
references.citation: Trends Biochem Sci. 2007 Mar;32(3):101-10
references.pmid: 17291766
references.citation: Proc Natl Acad Sci U S A. 2008 Oct 21;105(42):16137-41
references.pmid: 18852451
references.citation: Crit Rev Biochem Mol Biol. 2008 Jan-Feb;43(1):63-88
references.pmid: 18307109
references.citation: Biochemistry. 2011 Oct 25;50(42):8986-8
references.pmid: 21950770
references.citation: Chem Commun (Camb). 2007 Jan 28;(4):359-61
references.pmid: 17220970
references.citation: Nucleic Acids Res. 2001 Mar 1;29(5):1097-106
references.pmid: 11222759
references.citation: RNA. 2008 Jan;14(1):98-106
references.pmid: 18025251
references.citation: Nat Prod Rep. 2010 Jun;27(6):900-18
references.pmid: 20372740
references.citation: Biochemistry. 2009 Oct 27;48(42):10162-74
references.pmid: 19736993
references.citation: Annu Rev Microbiol. 2007;61:113-29
references.pmid: 17506685
references.citation: Science. 2004 Jan 2;303(5654):76-9
references.pmid: 14704425
references.citation: Chem Biol. 2009 Jan 30;16(1):70-81
references.pmid: 19171307
references.citation: J Antibiot (Tokyo). 1992 May;45(5):781-7
references.pmid: 1624380
references.citation: J Antibiot (Tokyo). 1992 Nov;45(11):1812-4
references.pmid: 1468993
references.citation: J Biol Chem. 1995 Jul 28;270(30):17758-64
references.pmid: 7629075
references.citation: Proc Natl Acad Sci U S A. 2011 Mar 8;108(10):3930-4
references.pmid: 21368151
references.citation: ACS Chem Biol. 2009 Oct 16;4(10):855-64
references.pmid: 19678698
references.citation: Proc Natl Acad Sci U S A. 2004 Aug 31;101(35):12870-5
references.pmid: 15317939
references.citation: Curr Opin Struct Biol. 2008 Dec;18(6):658-66
references.pmid: 19059104
references.citation: J Am Chem Soc. 2010 Mar 24;132(11):3953-64
references.pmid: 20184321
references.citation: F1000 Biol Rep. 2010 Jul 14;2:52
references.pmid: 21152342
references.citation: J Am Chem Soc. 2002 Sep 25;124(38):11270-1
references.pmid: 12236732
references.citation: Proc Natl Acad Sci U S A. 2010 May 11;107(19):8537-42
references.pmid: 20421508
references.citation: Chem Biol. 2000 Aug;7(8):623-42
references.pmid: 11048953
references.citation: Annu Rev Biochem. 2009;78:65-94
references.pmid: 19489722
references.citation: Chembiochem. 2003 Aug 4;4(8):768-73
references.pmid: 12898629
references.citation: J Org Chem. 2006 Dec 22;71(26):9561-71
references.pmid: 17168571
references.citation: Science. 2011 Apr 29;332(6029):604-7
references.pmid: 21415317
references.citation: Antimicrob Agents Chemother. 2006 Jul;50(7):2500-5
references.pmid: 16801432
references.citation: Mol Biosyst. 2007 Jan;3(1):60-74
references.pmid: 17216057
references.citation: Bioorg Chem. 2011 Dec;39(5-6):161-70
references.pmid: 21762947
references.citation: Amino Acids. 2008 Feb;34(2):187-93
references.pmid: 17334902
references.citation: Curr Opin Chem Biol. 2009 Feb;13(1):58-73
references.pmid: 19297239
references.citation: EMBO J. 2003 Dec 1;22(23):6214-24
references.pmid: 14633981
references.citation: Antimicrob Agents Chemother. 2000 Nov;44(11):3040-8
references.pmid: 11036020
references.citation: Curr Opin Struct Biol. 2010 Dec;20(6):684-92
references.pmid: 20951571
references.citation: Mol Biol Evol. 2007 Aug;24(8):1596-9
references.pmid: 17488738
references.citation: J Am Chem Soc. 2009 Apr 29;131(16):5946-55
references.pmid: 19338336
references.citation: Mol Biosyst. 2009 Jan;5(1):77-90
references.pmid: 19081934
references.citation: Angew Chem Int Ed Engl. 2000 Jan;39(2):393-395
references.pmid: 10649422
references.citation: Mol Biosyst. 2010 Jul;6(7):1180-5
references.pmid: 20473441
references.citation: Trends Biochem Sci. 2004 May;29(5):243-9
references.pmid: 15130560
references.citation: Chem Biol. 2003 Apr;10(4):301-11
references.pmid: 12725858
references.citation: Curr Opin Chem Biol. 2011 Apr;15(2):267-75
references.pmid: 21159543
references.citation: J Biol Chem. 2011 Sep 2;286(35):30245-30252
references.pmid: 21771780
references.citation: Nucleic Acids Res. 2004 Aug 02;32(13):4015-25
references.pmid: 15289575
references.citation: Arch Biochem Biophys. 1990 Apr;278(1):35-40
references.pmid: 2321967
references.citation: J Biol Chem. 2010 Feb 19;285(8):5792-801
references.pmid: 20007320
references.citation: Chembiochem. 2010 Mar 22;11(5):604-21
references.pmid: 20191656
references.citation: Nat Chem Biol. 2010 Aug;6(8):581-6
references.pmid: 20562876
references.citation: J Antibiot (Tokyo). 2007 Aug;60(8):492-503
references.pmid: 17827660
references.citation: Nat Prod Rep. 1999 Oct;16(5):589-96
references.pmid: 10584333
references.citation: J Antibiot (Tokyo). 1995 Oct;48(10):1191-3
references.pmid: 7490235
references.citation: J Biol Chem. 2005 Aug 12;280(32):29038-46
references.pmid: 15967800