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Structural analyses of NudT16-ADP-ribose complexes direct rational design of mutants with improved processing of poly(ADP-ribosyl)ated proteins.

ABSTRACT: ADP-ribosylation is a post-translational modification that occurs on chemically diverse amino acids, including aspartate, glutamate, lysine, arginine, serine and cysteine on proteins and is mediated by ADP-ribosyltransferases, including a subset commonly known as poly(ADP-ribose) polymerases. ADP-ribose can be conjugated to proteins singly as a monomer or in polymeric chains as poly(ADP-ribose). While ADP-ribosylation can be reversed by ADP-ribosylhydrolases, this protein modification can also be processed to phosphoribosylation by enzymes possessing phosphodiesterase activity, such as snake venom phosphodiesterase, mammalian ectonucleotide pyrophosphatase/phosphodiesterase 1, Escherichia coli RppH, Legionella pneumophila Sde and Homo sapiens NudT16 (HsNudT16). Our studies here sought to utilize X-ray crystallographic structures of HsNudT16 in complex with monomeric and dimeric ADP-ribose in identifying the active site for binding and processing free and protein-conjugated ADP-ribose into phosphoribose forms. These structural data guide rational design of mutants that widen the active site to better accommodate protein-conjugated ADP-ribose. We identified that several HsNudT16 mutants (?17, F36A, and F61S) have reduced activity for free ADP-ribose, similar processing ability against protein-conjugated mono(ADP-ribose), but improved catalytic efficiency for protein-conjugated poly(ADP-ribose). These HsNudT16 variants may, therefore, provide a novel tool to investigate different forms of ADP-ribose.

SUBMITTER: Thirawatananond P

PROVIDER: S-EPMC6459841 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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Structural analyses of NudT16-ADP-ribose complexes direct rational design of mutants with improved processing of poly(ADP-ribosyl)ated proteins.

Thirawatananond Puchong P McPherson Robert Lyle RL Malhi Jasmine J Nathan Sara S Lambrecht Michael J MJ Brichacek Matthew M Hergenrother Paul J PJ Leung Anthony K L AKL Gabelli Sandra B SB

Scientific reports 20190411 1

ADP-ribosylation is a post-translational modification that occurs on chemically diverse amino acids, including aspartate, glutamate, lysine, arginine, serine and cysteine on proteins and is mediated by ADP-ribosyltransferases, including a subset commonly known as poly(ADP-ribose) polymerases. ADP-ribose can be conjugated to proteins singly as a monomer or in polymeric chains as poly(ADP-ribose). While ADP-ribosylation can be reversed by ADP-ribosylhydrolases, this protein modification can also b ...[more]

PMID: 30976021

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Project description:Many of the long-term effects of cocaine on the brain's reward circuitry have been shown to be mediated by alterations in gene expression. Several chromatin modifications, including histone acetylation and methylation, have been implicated in this regulation, but the effect of other histone modifications remains poorly understood. Poly(ADP-ribose) polymerase-1 (PARP-1), a ubiquitous and abundant nuclear protein, catalyzes the synthesis of a negatively charged polymer called poly(ADP-ribose) or PAR on histones and other substrate proteins and forms transcriptional regulatory complexes with several other chromatin proteins. Here, we identify an essential role for PARP-1 in cocaine-induced molecular, neural, and behavioral plasticity. Repeated cocaine administration, including self-administration, increased global levels of PARP-1 and its mark PAR in mouse nucleus accumbens (NAc), a key brain reward region. Using PARP-1 inhibitors and viral-mediated gene transfer, we established that PARP-1induction in NAc mediates enhanced behavioral responses to cocaine, including increased self-administration of the drug. Using chromatin immunoprecipitation sequencing, we demonstrated a global, genome-wide enrichment of PARP-1 in NAc of cocaine-exposed mice and identified several PARP-1 target genes that could contribute to the lasting effects of cocaine. Specifically, we identified sidekick-1-important for synaptic connections during development-as a critical PARP-1 target gene involved in cocaine's behavioral effects as well as in its ability to induce dendritic spines on NAc neurons. These findings establish the involvement of PARP-1 and PARylation in the long-term actions of cocaine. c57bl/6 mice were given IP injections of chronic cocaine 20mg/kg once per day for 7 days and sacrificed 30 minutes after the final dose of cocaine. Control animals were given saline for 7 days and sacrificed 30 minutes after the final dose of saline. Nucleus accumbens (NAc) tissue was collected and then PARP-1 ChIP-seq was performed. Three sequencing replicates were performed on each group.

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Structural analyses of NudT16-ADP-ribose complexes direct rational design of mutants with improved processing of poly(ADP-ribosyl)ated proteins.

Publications

Structural analyses of NudT16-ADP-ribose complexes direct rational design of mutants with improved processing of poly(ADP-ribosyl)ated proteins.

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