PDBsum entry: 1ml9

Transferase

PDB id: 1ml9

Contents

Protein chain

260 a.a. *

Ligands

UNK ×6

Metals

_ZN ×3

Waters ×111

* Residue conservation analysis

PDB id:

1ml9

Name:

Transferase

Title:

Structure of the neurospora set domain protein dim-5, a histone lysine methyltransferase

Structure:

Histone h3 methyltransferase dim-5. Chain: a. Fragment: residues 17-318. Engineered: yes

Source:

Neurospora crassa. Organism_taxid: 5141. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Resolution:

1.98Å R-factor: 0.205 R-free: 0.258

Authors:

X.Zhang,H.Tamaru,S.I.Khan,J.R.Horton,L.J.Keefe,E.U.Selker, X.Cheng

Key ref:

X.Zhang et al. (2002). Structure of the Neurospora SET domain protein DIM-5, a histone H3 lysine methyltransferase. Cell, 111, 117-127. PubMed id: 12372305 DOI: 10.1016/S0092-8674(02)00999-6

Date:

30-Aug-02 Release date: 23-Oct-02

Protein chain

Q8X225  (DIM5_NEUCR) -  Histone-lysine N-methyltransferase, H3 lysine-9 specific dim-5

Seq: 331 a.a.

Struc: 260 a.a.

Enzyme reactions

• Enzyme class: E.C.2.1.1.43 - Histone-lysine N-methyltransferase.
• Reaction: S-adenosyl-L-methionine + L-lysine-[histone] = S-adenosyl-L-homocysteine + N₆-methyl-L-lysine-[histone]

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

 Gene Ontology (GO) functional annotation 

• Cellular component: nucleus (1 term)
• Biological process: chromatin modification (1 term)
• Biochemical function: protein binding (6 terms)

Added reference

DOI no: 10.1016/S0092-8674(02)00999-6

Cell 111:117-127 (2002)

PubMed id: 12372305

Structure of the Neurospora SET domain protein DIM-5, a histone H3 lysine methyltransferase.

X.Zhang, H.Tamaru, S.I.Khan, J.R.Horton, L.J.Keefe, E.U.Selker, X.Cheng.

ABSTRACT

AdoMet-dependent methylation of histones is part of the "histone code" that can profoundly influence gene expression. We describe the crystal structure of Neurospora DIM-5, a histone H3 lysine 9 methyltranferase (HKMT), determined at 1.98 A resolution, as well as results of biochemical characterization and site-directed mutagenesis of key residues. This SET domain protein bears no structural similarity to previously characterized AdoMet-dependent methyltransferases but includes notable features such as a triangular Zn3Cys9 zinc cluster in the pre-SET domain and a AdoMet binding site in the SET domain essential for methyl transfer. The structure suggests a mechanism for the methylation reaction and provides the structural basis for functional characterization of the HKMT family and the SET domain.

Selected figure(s)

Figure 3.
Figure 3. Enzymatic Properties of Recombinant DIM-5HKMT activity as functions of (A) temperature, (B) salt concentration, (C) pH, and (D) AdoMet crosslinking as a function of pH. The buffers used were 50 mM Na citrate for pH 5.0–6.0, MES for pH 6.0–6.5, HEPES for pH 7.0–7.5, Tris for pH 8.0–8.5, Bicine for pH 9.0, and glycine for pH 9.35–10.7. To rule out the potential inhibitory effect of Na citrate, both Na citrate and Mes are used for pH 6.0.(E) Activities of DIM-5 mutants with conservative point mutations. All mutant proteins were expressed to level similar to that of the wild-type, though some were less soluble, and all were monomeric, suggesting that none of the mutations caused gross aggregation of the protein. Various amounts of mutant enzymes were used, the activities were compared to that of serial dilutions of wild-type enzymes purified in the same way, and the specific activity of mutant proteins relative to wild-type was estimated. The activities shown are averages of at least two measurements.(F) Fluorographic results of an AdoMet crosslinking experiment at pH 8.0 are shown along with results of Coomassie staining to control for the amount of mutant protein tested.

Figure 6.
Figure 6. Metal Chelators Inhibit DIM-5 Activity(A) Analysis of zinc content of DIM-5 with and without EDTA treatment. DIM-5 protein was incubated with 20 mM EDTA for 2 days, at which time HKMT activity was no longer detectable. To remove zinc bound to EDTA, the protein was either dialyzed (Exp1) or subjected to gel filtration chromatography (Exp2) against 20 mM glycine (pH 9.8), 5% glycerol, 0.5 mM DTT, and 1 mM EDTA.(B) Purified DIM-5 protein (1 mg/ml in 20 mM glycine [pH 9.8], 5% glycerol) was incubated with various concentration of 1,10-phenanthroline or EDTA for 18 hr at 4°C. The enzyme was diluted 80-fold and assayed for HKMT activity under standard conditions, except that no DTT was present.(C) Fluorographic results of AdoMet crosslinking in the presence of EDTA.

The above figures are reprinted by permission from Cell Press: Cell (2002, 111, 117-127) copyright 2002.

Figures were selected by the author.