An engineered variant of SETD3 methyltransferase alters target specificity from histidine to lysine methylation
Abstract
Most characterized SET domain (SETD) proteins are protein lysine methyltransferases, but SETD3 was recently demonstrated to be a protein (i.e. actin) histidine-N3 methyltransferase. Human SETD3 shares a high structural homology with two known protein lysine methyltransferases—human SETD6 and the plant LSMT—but differs in the residues constituting the active site. In the SETD3 active site, Asn255 engages in a unique hydrogen-bonding interaction with the target histidine of actin that likely contributes to its >1300-fold greater catalytic efficiency (kcat/Km) on histidine than on lysine. In this work, we engineered active-site variants to switch the SETD3 target specificity from histidine to lysine. Substitution of Asn255 with phenylalanine (N255F), together with substitution of Trp273 with alanine (W273A), generated an active site mimicking that of known lysine methyltransferases. The doubly substituted SETD3 variant exhibited a 13-fold preference for lysine over histidine. We show, by means of X-ray crystallography, that the two target nitrogen atoms—the N3 atom of histidine and the terminal ϵ-amino nitrogen of lysine—occupy the same position and point toward and are within a short distance of the incoming methyl group of SAM for a direct methyl transfer during catalysis. In contrast, SETD3 and its Asn255 substituted derivatives did not methylate glutamine (another potentiallymore »
- Authors:
-
[2];
[1]
- Univ. of Texas M.D. Anderson Cancer Center, Houston, TX (United States)
- Stanford Univ., CA (United States)
- Publication Date:
- Research Org.:
- Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Org.:
- National Institutes of Health (NIH); Cancer Prevention and Research Institute of Texas (CPRIT)
- OSTI Identifier:
- 1615846
- Grant/Contract Number:
- GM114306; GM133051; RR160029
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Journal of Biological Chemistry
- Additional Journal Information:
- Journal Volume: 295; Journal Issue: 9; Journal ID: ISSN 0021-9258
- Publisher:
- American Society for Biochemistry and Molecular Biology
- Country of Publication:
- United States
- Language:
- ENGLISH
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; S-adenosylmethionine (SAM); protein methylation; structural biology; enzyme catalysis; enzyme kinetics; histidine; glutamine methylation; histidine methylation; lysine methylation
Citation Formats
Dai, Shaobo, Horton, John R., Wilkinson, Alex W., Gozani, Or, Zhang, Xing, and Cheng, Xiaodong. An engineered variant of SETD3 methyltransferase alters target specificity from histidine to lysine methylation. United States: N. p., 2020.
Web. doi:10.1074/jbc.RA119.012319.
Dai, Shaobo, Horton, John R., Wilkinson, Alex W., Gozani, Or, Zhang, Xing, & Cheng, Xiaodong. An engineered variant of SETD3 methyltransferase alters target specificity from histidine to lysine methylation. United States. https://doi.org/10.1074/jbc.RA119.012319
Dai, Shaobo, Horton, John R., Wilkinson, Alex W., Gozani, Or, Zhang, Xing, and Cheng, Xiaodong. Tue .
"An engineered variant of SETD3 methyltransferase alters target specificity from histidine to lysine methylation". United States. https://doi.org/10.1074/jbc.RA119.012319. https://www.osti.gov/servlets/purl/1615846.
@article{osti_1615846,
title = {An engineered variant of SETD3 methyltransferase alters target specificity from histidine to lysine methylation},
author = {Dai, Shaobo and Horton, John R. and Wilkinson, Alex W. and Gozani, Or and Zhang, Xing and Cheng, Xiaodong},
abstractNote = {Most characterized SET domain (SETD) proteins are protein lysine methyltransferases, but SETD3 was recently demonstrated to be a protein (i.e. actin) histidine-N3 methyltransferase. Human SETD3 shares a high structural homology with two known protein lysine methyltransferases—human SETD6 and the plant LSMT—but differs in the residues constituting the active site. In the SETD3 active site, Asn255 engages in a unique hydrogen-bonding interaction with the target histidine of actin that likely contributes to its >1300-fold greater catalytic efficiency (kcat/Km) on histidine than on lysine. In this work, we engineered active-site variants to switch the SETD3 target specificity from histidine to lysine. Substitution of Asn255 with phenylalanine (N255F), together with substitution of Trp273 with alanine (W273A), generated an active site mimicking that of known lysine methyltransferases. The doubly substituted SETD3 variant exhibited a 13-fold preference for lysine over histidine. We show, by means of X-ray crystallography, that the two target nitrogen atoms—the N3 atom of histidine and the terminal ϵ-amino nitrogen of lysine—occupy the same position and point toward and are within a short distance of the incoming methyl group of SAM for a direct methyl transfer during catalysis. In contrast, SETD3 and its Asn255 substituted derivatives did not methylate glutamine (another potentially methylated amino acid). However, the glutamine-containing peptide competed with the substrate peptide, and glutamine bound in the active site, but too far away from SAM to be methylated. Our results provide insight into the structural parameters defining the target amino acid specificity of SET enzymes.},
doi = {10.1074/jbc.RA119.012319},
journal = {Journal of Biological Chemistry},
number = 9,
volume = 295,
place = {United States},
year = {2020},
month = {1}
}
Free Publicly Available Full Text
Publisher's Version of Record
Other availability
Search WorldCat to find libraries that may hold this journal
Cited by: 1 work
Citation information provided by
Web of Science
Web of Science
Save to My Library
You must Sign In or Create an Account in order to save documents to your library.
Works referenced in this record:
KMT9 monomethylates histone H4 lysine 12 and controls proliferation of prostate cancer cells
journal, May 2019
- Metzger, Eric; Wang, Sheng; Urban, Sylvia
- Nature Structural & Molecular Biology, Vol. 26, Issue 5
Use of knowledge-based restraints in phenix.refine to improve macromolecular refinement at low resolution
journal, March 2012
- Headd, Jeffrey J.; Echols, Nathaniel; Afonine, Pavel V.
- Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4
SETD3 protein is the actin-specific histidine N-methyltransferase
journal, December 2018
- Kwiatkowski, Sebastian; Seliga, Agnieszka K.; Vertommen, Didier
- eLife, Vol. 7
Specificity Analysis-Based Identification of New Methylation Targets of the SET7/9 Protein Lysine Methyltransferase
journal, January 2011
- Dhayalan, Arunkumar; Kudithipudi, Srikanth; Rathert, Philipp
- Chemistry & Biology, Vol. 18, Issue 1
SET domain proteins modulate chromatin domains in eu- and heterochromatin
journal, January 1998
- Jenuwein, T.; Laible, G.; Dorn, R.
- Cellular and Molecular Life Sciences (CMLS), Vol. 54, Issue 1
DOT1L and H3K79 Methylation in Transcription and Genomic Stability
journal, February 2018
- Wood, Katherine; Tellier, Michael; Murphy, Shona
- Biomolecules, Vol. 8, Issue 1
SET8 Recognizes the Sequence RHRK 20 VLRDN within the N Terminus of Histone H4 and Mono-methylates Lysine 20
journal, June 2005
- Yin, Yinliang; Liu, Changdong; Tsai, Sau Na
- Journal of Biological Chemistry, Vol. 280, Issue 34
Analysis of the Substrate Specificity of the Dim-5 Histone Lysine Methyltransferase Using Peptide Arrays
journal, January 2008
- Rathert, Philipp; Zhang, Xing; Freund, Christian
- Chemistry & Biology, Vol. 15, Issue 1
Structural basis of SETD6-mediated regulation of the NF-kB network via methyl-lysine signaling
journal, April 2011
- Chang, Yanqi; Levy, Dan; Horton, John R.
- Nucleic Acids Research, Vol. 39, Issue 15
Coot model-building tools for molecular graphics
journal, November 2004
- Emsley, Paul; Cowtan, Kevin
- Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 12, p. 2126-2132
Enterovirus pathogenesis requires the host methyltransferase SETD3
journal, September 2019
- Diep, Jonathan; Ooi, Yaw Shin; Wilkinson, Alex W.
- Nature Microbiology, Vol. 4, Issue 12
Nonhistone Lysine Methylation in the Regulation of Cancer Pathways
journal, August 2016
- Carlson, Scott M.; Gozani, Or
- Cold Spring Harbor Perspectives in Medicine, Vol. 6, Issue 11
Expression, Purification, and Characterization of Recombinant Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Large SubunitNϵ-Methyltransferase
journal, October 1998
- Zheng, Qi; Simel, Erica J.; Klein, Patricia E.
- Protein Expression and Purification, Vol. 14, Issue 1
Structure of the Catalytic Domain of Human DOT1L, a Non-SET Domain Nucleosomal Histone Methyltransferase
journal, March 2003
- Min, Jinrong; Feng, Qin; Li, Zhizhong
- Cell, Vol. 112, Issue 5
Cloning and developmental expression of pea ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit N-methyltransferase
journal, January 1995
- Klein, Robert R.; Houtz, Robert L.
- Plant Molecular Biology, Vol. 27, Issue 2
Historical review: the field of protein methylation
journal, March 2007
- Paik, Woon Ki; Paik, David C.; Kim, Sangduk
- Trends in Biochemical Sciences, Vol. 32, Issue 3
Methyltransferase-Glo: a universal, bioluminescent and homogenous assay for monitoring all classes of methyltransferases
journal, March 2016
- Hsiao, Kevin; Zegzouti, Hicham; Goueli, Said A.
- Epigenomics, Vol. 8, Issue 3
Structural basis for the target specificity of actin histidine methyltransferase SETD3
journal, August 2019
- Dai, Shaobo; Horton, John R.; Woodcock, Clayton B.
- Nature Communications, Vol. 10, Issue 1
Structural insights into SETD3-mediated histidine methylation on β-actin
journal, February 2019
- Guo, Qiong; Liao, Shanhui; Kwiatkowski, Sebastian
- eLife, Vol. 8
Many paths to methyltransfer: a chronicle of convergence
journal, June 2003
- Schubert, Heidi L.; Blumenthal, Robert M.; Cheng, Xiaodong
- Trends in Biochemical Sciences, Vol. 28, Issue 6
SETD3 is an actin histidine methyltransferase that prevents primary dystocia
journal, December 2018
- Wilkinson, Alex W.; Diep, Jonathan; Dai, Shaobo
- Nature, Vol. 565, Issue 7739
Structure and Catalytic Mechanism of a SET Domain Protein Methyltransferase
journal, October 2002
- Trievel, Raymond C.; Beach, Bridgette M.; Dirk, Lynnette M. A.
- Cell, Vol. 111, Issue 1
The SET8 H4K20 protein lysine methyltransferase has a long recognition sequence covering seven amino acid residues
journal, November 2012
- Kudithipudi, Srikanth; Dhayalan, Arunkumar; Kebede, Adam Fiseha
- Biochimie, Vol. 94, Issue 11
Post-translational modifications and chromatin dynamics
journal, March 2019
- Tolsma, Thomas O.; Hansen, Jeffrey C.
- Essays in Biochemistry, Vol. 63, Issue 1
Structure of the Neurospora SET Domain Protein DIM-5, a Histone H3 Lysine Methyltransferase
journal, October 2002
- Zhang, Xing; Tamaru, Hisashi; Khan, Seema I.
- Cell, Vol. 111, Issue 1
Introduction to the multi-author review on methylation in cellular physiology
journal, June 2019
- Shechter, David
- Cellular and Molecular Life Sciences, Vol. 76, Issue 15
Lysine methylation signaling of non-histone proteins in the nucleus
journal, May 2019
- Levy, Dan
- Cellular and Molecular Life Sciences, Vol. 76, Issue 15
Human HemK2/KMT9/N6AMT1 is an active protein methyltransferase, but does not act on DNA in vitro, in the presence of Trm112
journal, September 2019
- Woodcock, Clayton B.; Yu, Dan; Zhang, Xing
- Cell Discovery, Vol. 5, Issue 1
Substrate Specificity of the HEMK2 Protein Glutamine Methyltransferase and Identification of Novel Substrates
journal, January 2016
- Kusevic, Denis; Kudithipudi, Srikanth; Jeltsch, Albert
- Journal of Biological Chemistry, Vol. 291, Issue 12
Protein lysine methyltransferase G9a acts on non-histone targets
journal, April 2008
- Rathert, Philipp; Dhayalan, Arunkumar; Murakami, Marie
- Nature Chemical Biology, Vol. 4, Issue 6
HemK2 protein, encoded on human chromosome 21, methylates translation termination factor eRF1
journal, June 2008
- Figaro, Sabine; Scrima, Nathalie; Buckingham, Richard H.
- FEBS Letters, Vol. 582, Issue 16
Multiparametric scaling of diffraction intensities
journal, April 2003
- Otwinowski, Zbyszek; Borek, Dominika; Majewski, Wladyslaw
- Acta Crystallographica Section A Foundations of Crystallography, Vol. 59, Issue 3
Structure of the Conserved Core of the Yeast Dot1p, a Nucleosomal Histone H3 Lysine 79 Methyltransferase
journal, July 2004
- Sawada, Ken; Yang, Zhe; Horton, John R.
- Journal of Biological Chemistry, Vol. 279, Issue 41
Phaser crystallographic software
journal, July 2007
- McCoy, Airlie J.; Grosse-Kunstleve, Ralf W.; Adams, Paul D.
- Journal of Applied Crystallography, Vol. 40, Issue 4
Lysine methylation of the NF-κB subunit RelA by SETD6 couples activity of the histone methyltransferase GLP at chromatin to tonic repression of NF-κB signaling
journal, December 2010
- Levy, Dan; Kuo, Alex J.; Chang, Yanqi
- Nature Immunology, Vol. 12, Issue 1
Regulation of chromatin structure by site-specific histone H3 methyltransferases
journal, August 2000
- Rea, Stephen; Eisenhaber, Frank; O'Carroll, Dónal
- Nature, Vol. 406, Issue 6796
Rubisco in complex with Rubisco large subunit methyltransferase
journal, February 2009
- Raunser, S.; Magnani, R.; Huang, Z.
- Proceedings of the National Academy of Sciences, Vol. 106, Issue 9