The dynamic conformational landscape of the protein methyltransferase SETD8
Abstract
Elucidating the conformational heterogeneity of proteins is essential for understanding protein function and developing exogenous ligands. With the rapid development of experimental and computational methods, it is of great interest to integrate these approaches to illuminate the conformational landscapes of target proteins. SETD8 is a protein lysine methyltransferase (PKMT), which functions in vivo via the methylation of histone and nonhistone targets. Utilizing covalent inhibitors and depleting native ligands to trap hidden conformational states, we obtained diverse X-ray structures of SETD8. These structures were used to seed distributed atomistic molecular dynamics simulations that generated a total of six milliseconds of trajectory data. Markov state models, built via an automated machine learning approach and corroborated experimentally, reveal how slow conformational motions and conformational states are relevant to catalysis. These findings provide molecular insight on enzymatic catalysis and allosteric mechanisms of a PKMT via its detailed conformational landscape.
Data availability
The molecular dynamics datasets generated and analyzed in this study are available via the Open Science Framework at https://osf.io/2h6p4.The code used for the generation and analysis of the molecular dynamics data is available via a Github repository at https://github.com/choderalab/SETD8-materials.PDB files: 6BOZ for BC-Inh1, 5W1Y for BC-Inh2, 4IJ8 for BC-SAM, and 5V2N for APO.
Article and author information
Author details
Funding
National Cancer Institute
- Jian Jin
- John D Chodera
- Minkui Luo
K. C. Wong Education Foundation
- Cheng Luo
Chinese Academy of Sciences
- Cheng Luo
National Natural Science Foundation of China
- Cheng Luo
the Tri-Institutional PhD Program in Chemical Biology
- Shi Chen
- Rafal P Wiewiora
Peer Reviewed Cancer Research Program of the Department of Defense
- Rafal P Wiewiora
AbbVie
- Peter J Brown
Bayer Pharma AG
- Peter J Brown
Boehringer Ingelheim
- Peter J Brown
Eshelman Institute for Innovation
- Peter J Brown
Genome Canada
- Peter J Brown
National Institute of General Medical Sciences
- Yujun George Zheng
- Jian Jin
- John D Chodera
- Minkui Luo
Innovative Medicines Initiative
- Peter J Brown
Canada Foundation for Innovation
- Peter J Brown
Janssen
- Peter J Brown
Merck & Co.
- Peter J Brown
Novartis Pharma AG
- Peter J Brown
Ontario Ministry of Economic Development and Innovation
- Peter J Brown
Pfizer
- Peter J Brown
São Paulo Research Foundation-FAPESP
- Peter J Brown
Takeda
- Hua Zou
- Robert J Skene
- Peter J Brown
the Wellcome Trust
- Peter J Brown
Eunice Kennedy Shriver National Institute of Child Health and Human Development
- Jian Jin
Starr Cancer Consortium
- John D Chodera
- Minkui Luo
MSKCC Functional Genomics Initiative
- John D Chodera
- Minkui Luo
The Sloan Kettering Institute
- Kyle A Beauchamp
- John D Chodera
- Minkui Luo
Mr. William H. Goodwin and Mrs. Alice Goodwin Commonwealth Foundation for Cancer Research, and the Experimental Therapeutics Center of Memorial Sloan Kettering Cancer Center
- Minkui Luo
Tri-Institutional Therapeutics Discovery Institute
- Minkui Luo
Louis V. Gerstner Young Investigator Award
- John D Chodera
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Sarel Jacob Fleishman, Weizmann Institute of Science, Israel
Publication history
- Received: January 22, 2019
- Accepted: May 8, 2019
- Accepted Manuscript published: May 13, 2019 (version 1)
- Version of Record published: June 17, 2019 (version 2)
Copyright
© 2019, Chen et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 5,629
- Page views
-
- 816
- Downloads
-
- 23
- Citations
Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Biochemistry and Chemical Biology
- Cell Biology
Dystroglycan (DG) requires extensive post-translational processing and O-glycosylation to function as a receptor for extracellular matrix (ECM) proteins containing laminin-G-like (LG) domains. Matriglycan is an elongated polysaccharide of alternating xylose (Xyl) and glucuronic acid (GlcA) that binds with high-affinity to ECM proteins with LG-domains and is uniquely synthesized on α-dystroglycan (α-DG) by like-acetylglucosaminyltransferase-1 (LARGE1). Defects in the post-translational processing or O-glycosylation of α-DG that result in a shorter form of matriglycan reduce the size of α-DG and decrease laminin binding, leading to various forms of muscular dystrophy. Previously, we demonstrated that Protein O-Mannose Kinase (POMK) is required for LARGE1 to generate full-length matriglycan on α-DG (~150-250 kDa) (Walimbe et al., 2020). Here, we show that LARGE1 can only synthesize a short, non-elongated form of matriglycan in mouse skeletal muscle that lacks the DG N-terminus (α-DGN), resulting in a ~100-125 kDa α-DG. This smaller form of α-DG binds laminin and maintains specific force but does not prevent muscle pathophysiology, including reduced force production after eccentric contractions or abnormalities in the neuromuscular junctions. Collectively, our study demonstrates that α-DGN, like POMK, is required for LARGE1 to extend matriglycan to its full mature length on α-DG and thus prevent muscle pathophysiology.
-
- Biochemistry and Chemical Biology
- Microbiology and Infectious Disease
The malaria parasite Plasmodium falciparum synthesizes significant amounts of phospholipids to meet the demands of replication within red blood cells. De novo phosphatidylcholine (PC) biosynthesis via the Kennedy pathway is essential, requiring choline that is primarily sourced from host serum lysophosphatidylcholine (lysoPC). LysoPC also acts as an environmental sensor to regulate parasite sexual differentiation. Despite these critical roles for host lysoPC, the enzyme(s) involved in its breakdown to free choline for PC synthesis are unknown. Here, we show that a parasite glycerophosphodiesterase (PfGDPD) is indispensable for blood stage parasite proliferation. Exogenous choline rescues growth of PfGDPD-null parasites, directly linking PfGDPD function to choline incorporation. Genetic ablation of PfGDPD reduces choline uptake from lysoPC, resulting in depletion of several PC species in the parasite, whilst purified PfGDPD releases choline from glycerophosphocholine in vitro. Our results identify PfGDPD as a choline-releasing glycerophosphodiesterase that mediates a critical step in PC biosynthesis and parasite survival.