EPR spectroscopy identifies Met and Lys residues that are essential for the interaction between the CusB N-terminal domain and metallochaperone CusF

Metallomics ◽  
2015 ◽  
Vol 7 (7) ◽  
pp. 1163-1172 ◽  
Author(s):  
Aviv Meir ◽  
Adi Natan ◽  
Yoni Moskovitz ◽  
Sharon Ruthstein
Keyword(s):  
Epr Spectroscopy ◽  
Terminal Domain ◽  
Lysine Residues

Methionine and lysine residues are important for preserving the structure of the CusB N-terminal domain and for the interaction with CusF.

scholarly journals EPR Spectroscopy of the C-terminal Domain of the M2 Protein from Influenza A Virus

2009 ◽  
Vol 96 (3) ◽  
pp. 432a
Author(s):  
Emily Brown ◽  
Phuong Nguyen ◽  
Kathleen P. Howard
Keyword(s):  
Influenza A Virus ◽  
Epr Spectroscopy ◽  
Influenza A ◽  
M2 Protein ◽  
Terminal Domain

scholarly journals The CRL4Cdt2 Ubiquitin Ligase Mediates the Proteolysis of Cyclin-Dependent Kinase Inhibitor Xic1 through a Direct Association with PCNA

2010 ◽  
Vol 30 (17) ◽  
pp. 4120-4133 ◽  
Author(s):  
Dong Hyun Kim ◽  
Varija N. Budhavarapu ◽  
Carlos R. Herrera ◽  
Hyung Wook Nam ◽  
Yu Sam Kim ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Proliferating Cell Nuclear Antigen ◽  
Kinase Inhibitor ◽  
Binding Domain ◽  
Nuclear Antigen ◽  
Cyclin Dependent Kinase ◽  
Cyclin Dependent Kinase Inhibitor ◽  
Terminal Domain ◽  
Lysine Residues ◽  
Proliferating Cell

ABSTRACT During DNA polymerase switching, the Xenopus laevis Cip/Kip-type cyclin-dependent kinase inhibitor Xic1 associates with trimeric proliferating cell nuclear antigen (PCNA) and is recruited to chromatin, where it is ubiquitinated and degraded. In this study, we show that the predominant E3 for Xic1 in the egg is the Cul4-DDB1-XCdt2 (Xenopus Cdt2) (CRL4Cdt2) ubiquitin ligase. The addition of full-length XCdt2 to the Xenopus extract promotes Xic1 turnover, while the N-terminal domain of XCdt2 (residues 1 to 400) cannot promote Xic1 turnover, despite its ability to bind both Xic1 and DDB1. Further analysis demonstrated that XCdt2 binds directly to PCNA through its C-terminal domain (residues 401 to 710), indicating that this interaction is important for promoting Xic1 turnover. We also identify the cis-acting sequences required for Xic1 binding to Cdt2. Xic1 binds to Cdt2 through two domains (residues 161 to 170 and 179 to 190) directly flanking the Xic1 PCNA binding domain (PIP box) but does not require PIP box sequences (residues 171 to 178). Similarly, human p21 binds to human Cdt2 through residues 156 to 161, adjacent to the p21 PIP box. In addition, we identify five lysine residues (K180, K182, K183, K188, and K193) immediately downstream of the Xic1 PIP box and within the second Cdt2 binding domain as critical sites for Xic1 ubiquitination. Our studies suggest a model in which both the CRL4Cdt2 E3- and PIP box-containing substrates, like Xic1, are recruited to chromatin through independent direct associations with PCNA.

scholarly journals Yeast PIAS-type Ull1/Siz1 Is Composed of SUMO Ligase and Regulatory Domains

2005 ◽  
Vol 280 (43) ◽  
pp. 35822-35828 ◽  
Author(s):  
Yoshimitsu Takahashi ◽  
Yoshiko Kikuchi
Keyword(s):  
Neck Region ◽  
Binding Motif ◽  
Conjugation System ◽  
Regulatory Domains ◽  
Terminal Domain ◽  
Sumo Ligase ◽  
Lysine Residues ◽  
Protein Instability ◽  
Two Hybrid

SUMO (small ubiquitin-like modifier)/Smt3 (suppressor of mif two) is a member of the ubiquitin-related protein family and is known to conjugate with many proteins. In the sumoylation pathway, SUMO/Smt3 is transferred to substrate lysine residues through the thioester cascade of E1 (activating enzyme) and E2 (conjugating enzyme), and E3 (SUMO ligase) functions as an adaptor between E2 and each substrate. Yeast Ull1 (ubiquitin-like protein ligase 1)/Siz1, a PIAS (protein inhibitor of activated STAT)-type SUMO ligase, modifies both cytoplasmic and nuclear proteins. In this paper, we performed a domain analysis of Ull1/Siz1 by constructing various deletion mutants. A novel conserved N-terminal domain, called PINIT, as well as the RING-like domain (SP-RING) were required for the SUMO ligase activity in the in vitro conjugation system and for interaction with Smt3 in an in vitro binding assay. The most distal N-terminal region, which contains a putative DNA-binding SAF-A/B, Acinus, and PIAS (SAP) motif, was not required for the ligase activity but was involved in nuclear localization. A strong SUMO-binding motif was identified, which interacted with Smt3 in the two-hybrid system but was not necessary for the ligase activity. The most distal C-terminal domain was important for stable localization at the bud neck region and thereby for the substrate recognition of septins. Furthermore, the C-terminal half conferred protein instability on Ull1/Siz1. Taken together, we conclude that the SP-RING and PINIT of Ull1/Siz1 are core domains of the SUMO ligase, and the other domains are regulatory for protein stability and subcellular localization.

scholarly journals A proline repeat domain in the Notch co-activator MAML1 is important for the p300-mediated acetylation of MAML1

2007 ◽  
Vol 404 (2) ◽  
pp. 289-298 ◽  
Author(s):  
Mariana Saint Just Ribeiro ◽  
Magnus L. Hansson ◽  
Annika E. Wallberg
Keyword(s):  
Target Genes ◽  
Repeat Motif ◽  
Terminal Domain ◽  
Evolutionarily Conserved ◽  
Repeat Domain ◽  
Intracellular Receptor ◽  
N Terminus ◽  
Lysine Residues ◽  
Receptor Domain

Ligand activation of Notch leads to the release of Notch IC (the intracellular receptor domain), which translocates to the nucleus and interacts with the DNA-binding protein CSL to control expression of specific target genes. In addition to ligand-mediated activation, Notch signalling can be further modulated by interactions of Notch IC with a number of other proteins. MAML1 has previously been shown to act co-operatively with the histone acetyltransferase p300 in Notch IC-mediated transcription. In the present study we show that the N-terminal domain of MAML1 directly interacts with both p300 and histones, and the p300–MAML1 complex specifically acetylates histone H3 and H4 tails in chromatin. Furthermore, p300 acetylates MAML1 and evolutionarily conserved lysine residues in the MAML1 N-terminus are direct substrates for p300-mediated acetylation. The N-terminal domain of MAML1 contains a proline repeat motif (PXPAAPAP) that was previously shown to be present in p53 and important for the p300–p53 interaction. We show that the MAML1 proline repeat motif interacts with p300 and enhances the activity of the MAML1 N-terminus in vivo. These findings suggest that the N-terminal domain of MAML1 plays an important role in Notch-regulated transcription, by direct interactions with Notch, p300 and histones.

scholarly journals Critical lysine residues within the overlooked N-terminal domain of human APE1 regulate its biological functions

2010 ◽  
Vol 38 (22) ◽  
pp. 8239-8256 ◽  
Author(s):  
Damiano Fantini ◽  
Carlo Vascotto ◽  
Daniela Marasco ◽  
Chiara D’Ambrosio ◽  
Milena Romanello ◽  
...  
Keyword(s):  
Biological Functions ◽  
Terminal Domain ◽  
Lysine Residues

scholarly journals HIV-1 Vif-mediated ubiquitination/degradation of APOBEC3G involves four critical lysine residues in its C-terminal domain

2009 ◽  
Vol 106 (46) ◽  
pp. 19539-19544 ◽  
Author(s):  
Y. Iwatani ◽  
D. S. B. Chan ◽  
L. Liu ◽  
H. Yoshii ◽  
J. Shibata ◽  
...  
Keyword(s):  
Terminal Domain ◽  
Lysine Residues ◽  
Hiv 1

scholarly journals Crosstalk between RNA Pol II C-Terminal Domain Acetylation and Phosphorylation via RPRD Proteins

2018 ◽  
Author(s):  
Ibraheem Ali ◽  
Diego Garrido Ruiz ◽  
Zuyao Ni ◽  
Jeffrey R. Johnson ◽  
Heng Zhang ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Isothermal Calorimetry ◽  
Peptide Binding ◽  
Rna Pol Ii ◽  
Post Translational Modifications ◽  
Pol Ii ◽  
Terminal Domain ◽  
Lysine Residues ◽  
Higher Eukaryotes ◽  
Heptad Repeats

SUMMARYPost-translational modifications of the RNA polymerase II C-terminal domain (CTD) coordinate the transcription cycle. Crosstalk between different modifications is poorly understood. Here, we show how acetylation of lysine residues at position 7 of characteristic heptad repeats (K7ac)—only found in higher eukaryotes—regulates phosphorylation of serines at position 5 (S5p), a conserved mark of polymerases initiating transcription. We identified the regulator of pre-mRNA domain-containing (RPRD) proteins as reader proteins of K7ac. K7ac enhanced CTD peptide binding to the CTD-interacting domain (CID) of RPRD1A and RPRD1B proteins in isothermal calorimetry and molecular modeling experiments. Deacetylase inhibitors increased K7ac- and decreased S5-phosphorylated polymerases, consistent with acetylation-dependent S5 dephosphorylation by an RPRD-associated S5 phosphatase. Consistent with this model, RPRD1B knockdown increased S5p, but enhanced K7ac, indicating RPRD proteins recruit K7 deacetylases, including HDAC1. We also report auto-regulatory crosstalk between K7ac and S5p via RPRD proteins and their interactions with acetyl- and phospho-eraser proteins.

scholarly journals Multiple C-Terminal Lysine Residues Target p53 for Ubiquitin-Proteasome-Mediated Degradation

2000 ◽  
Vol 20 (22) ◽  
pp. 8458-8467 ◽  
Author(s):  
Manuel S. Rodriguez ◽  
Joana M. P. Desterro ◽  
Sonia Lain ◽  
David P. Lane ◽  
Ronald T. Hay
Keyword(s):  
Transcriptional Activity ◽  
Ubiquitin Protein Ligase ◽  
Terminal Domain ◽  
Lysine Residues ◽  
Arginine Residues ◽  
Ubiquitin Proteasome ◽  
Ubiquitin Conjugation ◽  
Genotoxic Insult ◽  
E6 Protein

ABSTRACT In normal cells, p53 is maintained at a low level by ubiquitin-mediated proteolysis, but after genotoxic insult this process is inhibited and p53 levels rise dramatically. Ubiquitination of p53 requires the ubiquitin-activating enzyme Ubc5 as a ubiquitin conjugation enzyme and Mdm2, which acts as a ubiquitin protein ligase. In addition to the N-terminal region, which is required for interaction with Mdm2, the C-terminal domain of p53 modulates the susceptibility of p53 to Mdm2-mediated degradation. To analyze the role of the C-terminal domain in p53 ubiquitination, we have generated p53 molecules containing single and multiple lysine-to-arginine changes between residues 370 and 386. Although wild-type (WT) and mutant molecules show similar subcellular distributions, the mutants display a higher transcriptional activity than WT p53. Simultaneous mutation of lysine residues 370, 372, 373, 381, 382, and 386 to arginine residues (6KR p53 mutant) generates a p53 molecule with potent transcriptional activity that is resistant to Mdm2-induced degradation and is refractory to Mdm2-mediated ubiquitination. In contrast to WT p53, transcriptional activity directed by the 6KR p53 mutant fails to be negatively regulated by Mdm2. Those differences are also manifest in HeLa cells which express the human papillomavirus E6 protein, suggesting that p53 C-terminal lysine residues are also implicated in E6-AP-mediated ubiquitination. These data suggest that p53 C-terminal lysine residues are the main sites of ubiquitin ligation, which target p53 for proteasome-mediated degradation.

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