Structural modeling of g.Gly312Asp in the predicted Structure of MKRN3
There are currently no structures reported for the makorin protein family therefore it was
necessary to predict structure based on the separate domains identified from the literature.
Gene analysis suggested through nucleotide BLAST alignments that MKRN3 (ID Q13064)
comprises four zinc fingers1 ; ZNF1 95-122, ZNF2 238-265, ZNF3 311-365 and ZNF4 384-423,
with ZNF3 predicted to form a RING-type zinc finger domain whereas the other three form
C3H1-type 1 subclass of Zn finger domain (Fig. 1Bi).
The zinc finger sub families are defined according to motifs in which the position of cysteine
residues are crucial to metal ion binding, in Ring H2 the metal binding is Zn 2+ however other ZNF
motifs have been shown to coordinate other metals. Ring type zinc finger domains are defined
by
the
motif
Cys-Xz-Cys-X(9-39)-Cys-X(l-3)-His-X(2-3)-Cys-X2-Cys-X(4-48)-Cys-X2-Cys
with
two
unstructured loops of indeterminate length. 2-3
Structural model for ZNF3 of MKRN3
The mutation at 312 of the mature protein sequence sits at the start of a RING type zinc finger
region therefore protein BLAST4 alignments specifically for that region identified several
structural hits (supplementary table 1). A Phyre24 search also carried out identified of the top
twenty hits, structures aligned best with the ZNF1 and ZNF3 region (Fig. 1B, Supplementary
table 2) the top hits for ZNF3 from Phyre2 matched Human Ubiquitin Ligase NIRF (PDB
ID_1Z6U) whereas the top hit (with most coverage) from protein-BLAST (with also best pairwise
score) was for the protein Rad18 Ubiquitin Ligase Ring Domain Structure5 (PDB ID_2Y43), which
aligns in region 306-394 of MKRN3 (Fig. 1Bii). The structure of the ZNF3 domain was therefore
modelled independently with both these PDB in order to best explore a range of possible
structure orientations for MKRN3 ZNF3.
1
Jong, M.T., Carey, A.H., Caldwell, K.A., Lau, M.H., Handel, M.A., Driscoll, D.J., Stewart, C.L.,
Rinchik, E.M. & Nicholls, R.D. (1999) Imprinting of a RING zinc-finger encoding gene in the mouse
chromosome region homologous to the Prader-Willi syndrome genetic region. Hum Mol Genet 8, 795803.
2
Borden, K.L. & Freemont, P.S. (1996) The RING finger domain: a recent example of a sequencestructure family. Curr Opin Struct Biol 6, 395-401.
3
Lorick, K.L., Jensen, J.P., Fang, S., Ong, A.M., Hatakeyama, S. & Weissman, A.M. (1999) RING
fingers mediate ubiquitin-conjugating enzyme (E2)-dependent ubiquitination. Proc Natl Acad Sci U S A
96, 11364-11369.
4
Altschul, S.F., Madden, T.L., Schaffer, A.A., Zhang, J., Zhang, Z., Miller, W. & Lipman, D.J. (1997)
Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res
25, 3389-3402.
5
Huang, A., Hibbert, R.G., de Jong, R.N., Das, D., Sixma, T.K. & Boelens, R. (2011) Symmetry and
asymmetry of the RING-RING dimer of Rad18. J Mol Biol 410, 424-435.