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Human Protein-l-isoaspartate O-Methyltransferase Domain-Containing Protein 1 (PCMTD1) Associates with Cullin-RING Ligase Proteins

Warmack, Rebeccah A. and Pang, Eric Z. and Peluso, Esther and Lowenson, Jonathan D. and Ong, Joseph Y. and Torres, Jorge Z. and Clarke, Steven G. (2022) Human Protein-l-isoaspartate O-Methyltransferase Domain-Containing Protein 1 (PCMTD1) Associates with Cullin-RING Ligase Proteins. Biochemistry, 61 (10). pp. 879-894. ISSN 0006-2960. doi:10.1021/acs.biochem.2c00130.

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The spontaneous l-isoaspartate protein modification has been observed to negatively affect protein function. However, this modification can be reversed in many proteins in reactions initiated by the protein-l-isoaspartyl (d-aspartyl) O-methyltransferase (PCMT1). It has been hypothesized that an additional mechanism exists in which l-isoaspartate-damaged proteins are recognized and proteolytically degraded. Herein, we describe the protein-l-isoaspartate O-methyltransferase domain-containing protein 1 (PCMTD1) as a putative E3 ubiquitin ligase substrate adaptor protein. The N-terminal domain of PCMTD1 contains l-isoaspartate and S-adenosylmethionine (AdoMet) binding motifs similar to those in PCMT1. This protein also has a C-terminal domain containing suppressor of cytokine signaling (SOCS) box ubiquitin ligase recruitment motifs found in substrate receptor proteins of the Cullin-RING E3 ubiquitin ligases. We demonstrate specific PCMTD1 binding to the canonical methyltransferase cofactor S-adenosylmethionine (AdoMet). Strikingly, while PCMTD1 is able to bind AdoMet, it does not demonstrate any l-isoaspartyl methyltransferase activity under the conditions tested here. However, this protein is able to associate with the Cullin-RING proteins Elongins B and C and Cul5 in vitro and in human cells. The previously uncharacterized PCMTD1 protein may therefore provide an alternate maintenance pathway for modified proteins in mammalian cells by acting as an E3 ubiquitin ligase adaptor protein.

Item Type:Article
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URLURL TypeDescription
Warmack, Rebeccah A.0000-0002-9612-0511
Torres, Jorge Z.0000-0002-2158-889X
Clarke, Steven G.0000-0002-7303-6632
Additional Information:© 2022 American Chemical Society. Received 7 March 2022. Revised 15 April 2022. Published online 29 April 2022. Published in issue 17 May 2022. The authors would like to acknowledge Dr. Mark Arbing and the UCLA DOE Protein Expression Core for aid in the design and synthesis of the plasmids used within this study. The authors would also like to thank Austin Gable, Georgiana Salant, Andrea Hadjikyriacou, Kennen MacKay, Calvin Lin, and Dylan Valencia for their contributions at the initial stages of this project. This work was supported by the National Science Foundation grant MCB-1714569 (to S.G.C.), the National Institutes of Health grant R35GM139539 (to J.Z.T), and by funds from the UCLA Academic Senate Faculty Research Program, the Life Extension Foundation, Inc., and the Elizabeth and Thomas Plott Chair in Gerontology of the UCLA Longevity Center (to S.G.C.). R.A.W. and J.Y.O. were supported by the National Institutes of Health Ruth L. Kirschstein National Research Service Award GM007185. E.Z.P. was supported by the NIH NIGMS-funded predoctoral fellowship (T32GM136614). J.Y.O. was supported by a National Science Foundation Graduate Research Fellowship DGE-1650604 and by a UCLA Whitcome Predoctoral Fellowship in Molecular Biology. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The authors declare no competing financial interest. This study was performed in accordance with animal use protocols approved by the UCLA Animal Research Committee (Protocol 1993-109-64). Mice were scheduled to be euthanized if they met any early removal criteria (kyphosis, lack of grooming behavior). However, this did not occur with any of the animals in our study. Figures and tables were arranged in Adobe Illustrator. Signals from fluorographs were quantified by densitometry using ImageJ. (53) All data described in the manuscript are contained within the manuscript. Additional data are available upon request. Accession Codes: PCMT1: P22061-1, PCMTD1: Q96MG8-1, PCMTD2: Q9NV79-1, EloB: Q15370-1, EloC: Q15369-2, Cul5: Q93034, Rbx2: Q9WTZ1.
Funding AgencyGrant Number
Life Extension FoundationUNSPECIFIED
NIH Predoctoral FellowshipGM007185
NIH Predoctoral FellowshipT32GM136614
NSF Graduate Research FellowshipDGE-1650604
Subject Keywords:Alcohols, Genetics, Protein dynamics, Rodent models
Issue or Number:10
Record Number:CaltechAUTHORS:20220721-8102000
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Official Citation:Human Protein-l-isoaspartate O-Methyltransferase Domain-Containing Protein 1 (PCMTD1) Associates with Cullin-RING Ligase Proteins Rebeccah A. Warmack, Eric Z. Pang, Esther Peluso, Jonathan D. Lowenson, Joseph Y. Ong, Jorge Z. Torres, and Steven G. Clarke Biochemistry 2022 61 (10), 879-894 DOI: 10.1021/acs.biochem.2c00130
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:115723
Deposited By: George Porter
Deposited On:22 Jul 2022 15:36
Last Modified:22 Jul 2022 15:36

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