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bioRxiv · 10.1101/776112

Structural Basis of Protein Arginine Methyltransferase Activation by a Catalytically Dead Homolog (Prozyme)

Hashimoto, H.·Kafkova, L.·Raczkowski, A.·Jordan, K. D.·Read, L. K.·Debler, E. W.
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Abstract

Prozymes are pseudoenzymes that stimulate the function of weakly active enzymes through complex formation. The major Trypanosoma brucei protein arginine methyltransferase, TbPRMT1 enzyme (ENZ), requires TbPRMT1 prozyme (PRO) to form an active heterotetrameric complex. Here we present the x-ray crystal structure of the ENZ-{Delta}52PRO tetrameric complex with the cofactor product S-adenosyl-L-homocysteine (AdoHcy) at 2.4 [A] resolution. The individual ENZ and PRO units adopt the highly-conserved PRMT domain architecture and form an antiparallel heterodimer that corresponds to the canonical homodimer observed in all previously reported PRMTs. In turn, two such heterodimers assemble into a tetramer both in the crystal and in solution with twofold rotational symmetry. ENZ is unstable in absence of PRO and incapable of forming a homodimer due to a steric clash of a non-conserved tyrosine within the dimerization arm, rationalizing why PRO is required to complement ENZ to form a PRMT dimer that is necessary, but not sufficient for PRMT activity. The PRO structure deviates from other, active PRTMs in that it lacks the conserved {eta}2 310-helix within the Rossmann fold, abolishing co-factor binding. In addition to its chaperone function for ENZ, PRO substantially contributes to substrate binding. Heterotetramerization is required for catalysis, since heterodimeric ENZ-PRO mutants lack binding affinity and methyltransferase activity towards the substrate protein TbRGG1. Together, we provide a structural basis for PRMT1 ENZ activation by PRO heterotetramer formation, which is conserved across all kinetoplastids, and describe a chaperone function of the PRMT1 prozyme, which represents a novel mode of PRMT regulation.\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC=\"FIGDIR/small/776112v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (56K):\norg.highwire.dtl.DTLVardef@3d970dorg.highwire.dtl.DTLVardef@187f03forg.highwire.dtl.DTLVardef@1adee39org.highwire.dtl.DTLVardef@1f30f34_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG HighlightsO_LIThe crystal structure of Trypanosoma brucei protein arginine methyltransferase 1 (PRMT1) is reported\nC_LIO_LITwo enzyme-prozyme heterodimers form a stable heterotetramer essential for catalysis\nC_LIO_LIThe catalytically dead prozyme lacks elements essential for AdoMet binding\nC_LIO_LIThe enzyme alone is unstable and cannot form a canonical dimer due to steric clashes\nC_LIO_LIT. brucei PRMT1 prozyme adopts a chaperone function conserved across kinetoplastids\nC_LI

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Hashimoto, H., Kafkova, L., Raczkowski, A., Jordan, K. D., Read, L. K., Debler, E. W.. 2019-09-19. Structural Basis of Protein Arginine Methyltransferase Activation by a Catalytically Dead Homolog (Prozyme). https://doi.org/10.1101/776112

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