A glutamate/aspartate switch controls product specificity in a protein arginine methyltransferase

Journal Article · Mon Feb 08 00:00:00 EST 2016 · Proceedings of the National Academy of Sciences of the United States of America

DOI:https://doi.org/10.1073/pnas.1525783113· OSTI ID:1258672

Debler, Erik W. ^[1]; Jain, Kanishk ^[2]; Warmack, Rebeccah A. ^[2]; Feng, You ^[2]; Clarke, Steven G. ^[2]; Blobel, Günter ^[1]; Stavropoulos, Pete ^[1]

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1. Rockefeller Univ., New York, NY (United States)
2. Univ. of California, Los Angeles, CA (United States)

Trypanosoma brucei PRMT7 (TbPRMT7) is a protein arginine methyltransferase (PRMT) that strictly monomethylates various substrates, thus classifying it as a type III PRMT. However, the molecular basis of its unique product specificity has remained elusive. Here, in this work, we present the structure of TbPRMT7 in complex with its cofactor product S-adenosyl-l-homocysteine (AdoHcy) at 2.8 Å resolution and identify a glutamate residue critical for its monomethylation behavior. TbPRMT7 comprises the conserved methyltransferase and β-barrel domains, an N-terminal extension, and a dimerization arm. The active site at the interface of the N-terminal extension, methyltransferase, and β-barrel domains is stabilized by the dimerization arm of the neighboring protomer, providing a structural basis for dimerization as a prerequisite for catalytic activity. Mutagenesis of active-site residues highlights the importance of Glu181, the second of the two invariant glutamate residues of the double E loop that coordinate the target arginine in substrate peptides/proteins and that increase its nucleophilicity. Strikingly, mutation of Glu181 to aspartate converts TbPRMT7 into a type I PRMT, producing asymmetric dimethylarginine (ADMA). Isothermal titration calorimetry (ITC) using a histone H4 peptide showed that the Glu181Asp mutant has markedly increased affinity for monomethylated peptide with respect to the WT, suggesting that the enlarged active site can favorably accommodate monomethylated peptide and provide sufficient space for ADMA formation. In conclusion, these findings yield valuable insights into the product specificity and the catalytic mechanism of protein arginine methyltransferases and have important implications for the rational (re)design of PRMTs.

• Research Organization: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
• Sponsoring Organization: National Cancer Institute (NCI); National Institute of General Medical Sciences; National Institutes of Health (NIH); Ruth L. Kirschstein National Service Award
• OSTI ID: 1258672
• Alternate ID(s): OSTI ID: 1354247
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America, Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Issue: 8 Vol. 113; ISSN 0027-8424
• Publisher: National Academy of Sciences
• Country of Publication: United States
• Language: ENGLISH

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
PRMT
crystal structure
enzyme catalysis
epigenetics
histone methylation