The structure of the C4C4 ring finger of human NOT4 reveals features distinct from those of C3HC4 RING fingers.
The NOT4 protein is a component of the CCR4.NOT complex, a global regulator of RNA polymerase II transcription. Human NOT4 (hNOT4) contains a RING finger motif of the C(4)C(4) type. We expressed and purified the N-terminal region of hNOT4 (residues 1-78) encompassing the RING finger motif and determined the solution structure by heteronuclear NMR. NMR experiments using a (113)Cd-substituted hNOT4 RING finger showed that two metal ions are bound through cysteine residues in a cross-brace manner. The three-dimensional structure of the hNOT4 RING finger was refined with root mean square deviation values of 0.58 +/- 0.13 A for the backbone atoms and 1.08 +/- 0.12 A for heavy atoms. The hNOT4 RING finger consists of an alpha-helix and three long loops that are stabilized by zinc coordination. The overall folding of the hNOT4 RING finger is similar to that of the C(3)HC(4) RING fingers. The relative orientation of the two zinc-chelating loops and the alpha-helix is well conserved. However, for the other regions, the secondary structural elements are distinct.
Hanzawa
H
H
Bijvoet Center for Biomolecular Research, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
de Ruwe
M J
MJ
Albert
T K
TK
van Der Vliet
P C
PC
Timmers
H T
HT
Boelens
R
R
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001665:Binding Sites; D003545:Cysteine / Q000737:chemistry; D004268:DNA-Binding Proteins; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000737:chemistry*; D014158:Transcription, Genetic; D044767:Ubiquitin-Protein Ligases; D016335:Zinc Fingers
D004268:DNA-Binding Proteins; C539882:NR4A2 protein, human; D057126:Nuclear Receptor Subfamily 4, Group A, Member 2; D012097:Repressor Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C086923:MOT2 protein, S cerevisiae; D044767:Ubiquitin-Protein Ligases; D003545:Cysteine