title: Distributive Processing by the Iron(II)/α-Ketoglutarate-Dependent Catalytic Domains of the TET Enzymes Is Consistent with Epigenetic Roles for Oxidized 5-Methylcytosine Bases.
abstract: The ten-eleven translocation (TET) proteins catalyze oxidation of 5-methylcytosine ((5m)C) residues in nucleic acids to 5-hydroxymethylcytosine ((5hm)C), 5-formylcytosine ((5f)C), and 5-carboxycytosine ((5ca)C). These nucleotide bases have been implicated as intermediates on the path to active demethylation, but recent reports have suggested that they might have specific regulatory roles in their own right. In this study, we present kinetic evidence showing that the catalytic domains (CDs) of TET2 and TET1 from mouse and their homologue from Naegleria gruberi, the full-length protein NgTET1, are distributive in both chemical and physical senses, as they carry out successive oxidations of a single (5m)C and multiple (5m)C residues along a polymethylated DNA substrate. We present data showing that the enzyme neither retains (5hm)C/(5f)C intermediates of preceding oxidations nor slides along a DNA substrate (without releasing it) to process an adjacent (5m)C residue. These findings contradict a recent report by Crawford et al. ( J. Am. Chem. Soc. 2016 , 138 , 730 ) claiming that oxidation of (5m)C by CD of mouse TET2 is chemically processive (iterative). We further elaborate that this distributive mechanism is maintained for TETs in two evolutionarily distant homologues and posit that this mode of function allows the introduction of (5m)C forms as epigenetic markers along the DNA.
authors.affiliation: New England Biolabs, Inc. , 240 County Road, Ipswich, Massachusetts 01938, United States.
authors.forename: Esta
authors.initials: E
authors.lastname: Tamanaha
authors.affiliation: New England Biolabs, Inc. , 240 County Road, Ipswich, Massachusetts 01938, United States.
authors.forename: Shengxi
authors.initials: S
authors.lastname: Guan
authors.affiliation: New England Biolabs, Inc. , 240 County Road, Ipswich, Massachusetts 01938, United States.
authors.forename: Katherine
authors.initials: K
authors.lastname: Marks
authors.affiliation: New England Biolabs, Inc. , 240 County Road, Ipswich, Massachusetts 01938, United States.
authors.forename: Lana
authors.initials: L
authors.lastname: Saleh
mesh_terms: D044503:5-Methylcytosine / Q000378:metabolism*; D000818:Animals; D020134:Catalytic Domain; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D049308:Dioxygenases; D044127:Epigenesis, Genetic; D007501:Iron / Q000378:metabolism*; D007656:Ketoglutaric Acids / Q000378:metabolism*; D051379:Mice; D009253:Naegleria / Q000201:enzymology; D010084:Oxidation-Reduction; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D007656:Ketoglutaric Acids; D011518:Proto-Oncogene Proteins; C540839:TET1 protein, mouse; D044503:5-Methylcytosine; D007501:Iron; D049308:Dioxygenases; C551914:Tet2 protein, mouse