title: Preparation of Human Nuclear RNA m⁶A Methyltransferases and Demethylases and Biochemical Characterization of Their Catalytic Activity.
abstract: N(6)-Methyladenosine (m(6)A) represents the most prevalent internal modification in messenger and long noncoding RNAs. There has been a surge of interest toward understanding the biological significance of m(6)A modification. In this chapter, we describe the methods for biochemically studying the recently uncovered m(6)A methyltransferases (METTL3 and METTL14) and demethylases (FTO and ALKBH5). How to express these proteins, perform their biochemistry reactions against various RNA probes, and characterize the methylation and demethylation activity will be discussed.
authors.affiliation: Department of Chemistry, The University of Chicago, Chicago, Illinois, USA; Institute for Biophysical Dynamics, The University of Chicago, Chicago, Illinois, USA; Howard Hughes Medical Institute, The University of Chicago, Chicago, Illinois, USA.
authors.forename: Jianzhao
authors.initials: J
authors.lastname: Liu
authors.affiliation: Department of Chemistry, The University of Chicago, Chicago, Illinois, USA; Institute for Biophysical Dynamics, The University of Chicago, Chicago, Illinois, USA; Howard Hughes Medical Institute, The University of Chicago, Chicago, Illinois, USA.
authors.forename: Yanan
authors.initials: Y
authors.lastname: Yue
authors.affiliation: Department of Chemistry, The University of Chicago, Chicago, Illinois, USA; Institute for Biophysical Dynamics, The University of Chicago, Chicago, Illinois, USA; Howard Hughes Medical Institute, The University of Chicago, Chicago, Illinois, USA; Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, Illinois, USA. Electronic address: chuanhe@uchicago.edu.
authors.forename: Chuan
authors.initials: C
authors.lastname: He
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000235:genetics; D000071502:AlkB Homolog 5, RNA Demethylase; D000071516:Alpha-Ketoglutarate-Dependent Dioxygenase FTO; D002384:Catalysis; D049308:Dioxygenases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D008780:Methyltransferases / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D062085:RNA, Long Noncoding / Q000737:chemistry / Q000235:genetics*; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics; D015138:RNA, Nuclear / Q000737:chemistry* / Q000235:genetics
chemical_list: D008565:Membrane Proteins; D011506:Proteins; D062085:RNA, Long Noncoding; D012333:RNA, Messenger; D015138:RNA, Nuclear; C010223:N-methyladenosine; D049308:Dioxygenases; C559920:ALKBH5 protein, human; D000071502:AlkB Homolog 5, RNA Demethylase; D000071516:Alpha-Ketoglutarate-Dependent Dioxygenase FTO; C526492:FTO protein, human; C587355:METTL14 protein, human; D008780:Methyltransferases; C468275:METTL3 protein, human; D000241:Adenosine
keywords: ALKBH5; Demethylation activity; FTO; METTL3–METTL14; Methylation activity; N(6)-methyladenosine; m(6)A demethylase; m(6)A methyltransferase