Abstract
The ASHH2 CW domain is responsible for recognizing the methylation state at lysine 4 of histone 3 N-terminal tails and implicated in the recruitment of the ASHH2 methyltransferase enzyme correctly to the histones. The ASHH2 CW domain binds H3 lysine motifs that can be either mono-, di-, or tri-methylated [ARTK(meX)QTAR, where X denotes the number of methylations], but binds strongest to monomethylated instances (Kd values reported in the range of 1 µm to 500 nM). Hoppmann et al. published the uncomplexed NMR structure of an ASHH2 CW domain in 2011. Here we document the assignment of a shortened ASHH2 CW construct, CW42, with similar binding affinity and better expression yields than the one used to solve the uncomplexed structure. We also perform 1H–15N HSQC-monitored titrations that document at which protein–peptide ratios the complex is saturated. Backbone resonance assignments are presented for this shortened ASHH2 CW domain alone and bound to an H3 histone tail mimicking peptide monomethylated on lysine 4 (ARTK(me1)QTAR). Likewise, the assignment was also performed for the protein in complex with the dimethylated (ARTK(me2)QTAR) and trimethylated (ARTK(me3)QTAR) peptide. Overall, these two latter situations displayed a similar perturbation of shifts as the mono-methylated instance. In the case of the monomethylated histone tail mimic, side-chain assignment of CW42 in this complex was performed and reported in addition to backbone assignment, in preparation of a future solution structure determination and dynamics characterization of the CW42–ARTK(me1)QTAR complex.
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20 September 2021
ORCID id has been removed for author Olena Dobrovolska in the article.
Abbreviations
- H3:
-
Histone 3
- H3K4me1:
-
ARTK(me1)QTAR
- H3K4me2:
-
ARTK(me2)QTAR
- H3K4me3:
-
ARTK(me3)QTAR
- PTM:
-
Post-translational modification
References
Andrews FH et al (2016) Multivalent chromatin engagement and inter-domain crosstalk regulate MORC3 ATPase. Cell Rep 16(12):3195–3207
Furukawa A et al (2016) Quantitative analysis of protein-ligand interactions by NMR. Prog Nucl Magn Reson Spectrosc 96:47–57
Gasser SM et al (1998) The genetics of epigenetics. Cell Mol Life Sci 54(1):1–5
Grewal SIS, Jia ST (2007) Heterochromatin revisited. Nat Rev Genet 8(1):35–46
Grini PE et al (2009) The ASH1 HOMOLOG 2 (ASHH2) histone H3 methyltransferase is required for ovule and anther development in Arabidopsis. PLoS ONE 4(11):e7817
Hall TA (1999) BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. In: Nucleic acids symposium series. Information Retrieval Ltd., London, pp c1979–c2000
He FH et al (2010) Structural insight into the zinc finger CW domain as a histone modification reader. Structure 18(9):1127–1139
Hoppmann V et al (2011) The CW domain, a new histone recognition module in chromatin proteins. EMBO J 30(10):1939–1952
Keller RLJ (2005) Optimizing the process of nuclear magnetic resonance spectrum analysis and computer aided resonance assignment. Doctoral dissertation, ETH Zurich
Kim SY et al (2005) Establishment of the vernalization-responsive, winter-annual habit in Arabidopsis requires a putative histone H3 methyl transferase. Plant Cell 17(12):3301–3310
Mellor J (2006) Dynamic nucleosomes and gene transcription. Trends Genet 22(6):320–329
Michaels SD, Amasino RM (1999) Flowering Locus C encodes a novel MADS domain protein that acts as a repressor of flowering. Plant Cell 11(5):949–956
Morris SA et al (2005) Histone H3 k36 methylation is associated with transcription elongation in Schizosaccharomyces pombe. Eukaryot Cell 4(8):1446–1454
Williamson MP (2013) Using chemical shift perturbation to characterise ligand binding. Prog Nucl Magn Reson Spectrosc 73:1–16
Zhao Z et al (2005) Prevention of early flowering by expression of Flowering Locus C requires methylation of histone H3 K36. Nat Cell Biol 7(12):1256–1260
Acknowledgements
The authors would like to acknowledge the Norwegian NMR Platform (NNP) for support in the form of instruments access and experimental time (Grant 226244/F50). The authors wish to thank the NNP Bergen Node staff for facilitating the research. The authors also wish to thank Drs. V. De Marco, E. Christodoulou, and S. J. Gamblin, NIMR/MRC, London, for their advice and generous support for construction of CW42.
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12104_2018_9811_MOESM1_ESM.tif
Superimposition of the 1H–15N HSQC spectra for CW42 (current study, red) and previous CWs construct (Hoppmann et al., green) illustrating the difference between them and the need for a new assignment of CW42. (TIF 3739 KB)
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Dobrovolska, O., Bril’kov, M., Ødegård-Fougner, Ø. et al. 1H, 13C, and 15N resonance assignments of CW domain of the N-methyltransferase ASHH2 free and bound to the mono-, di- and tri-methylated histone H3 tail peptides. Biomol NMR Assign 12, 215–220 (2018). https://doi.org/10.1007/s12104-018-9811-x
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DOI: https://doi.org/10.1007/s12104-018-9811-x