Pub Date : 2023-02-01DOI: 10.1007/s12104-023-10118-6
Stéphane Thore, Sébastien Fribourg, Cameron D. Mackereth
The initial pre-mRNA transcript in eukaryotes is processed by a large multi-protein complex in order to correctly cleave the 3’ end, and to subsequently add the polyadenosine tail. This cleavage and polyadenylation specificity factor (CPSF) is composed of separate subunits, with structural information available for both isolated subunits and also larger assembled complexes. Nevertheless, certain key components of CPSF still lack high-resolution atomic data. One such region is the heterodimer formed between the first and second C-terminal domains of the endonuclease CPSF73, with those from the catalytically inactive CPSF100. Here we report the backbone and sidechain resonance assignments of a minimal C-terminal heterodimer of CPSF73–CPSF100 derived from the parasite Encephalitozoon cuniculi. The assignment process used several amino-acid specific labeling strategies, and the chemical shift values allow for secondary structure prediction.
{"title":"1H, 15N and 13C resonance assignments of a minimal CPSF73-CPSF100 C-terminal heterodimer","authors":"Stéphane Thore, Sébastien Fribourg, Cameron D. Mackereth","doi":"10.1007/s12104-023-10118-6","DOIUrl":"10.1007/s12104-023-10118-6","url":null,"abstract":"<div><p>The initial pre-mRNA transcript in eukaryotes is processed by a large multi-protein complex in order to correctly cleave the 3’ end, and to subsequently add the polyadenosine tail. This cleavage and polyadenylation specificity factor (CPSF) is composed of separate subunits, with structural information available for both isolated subunits and also larger assembled complexes. Nevertheless, certain key components of CPSF still lack high-resolution atomic data. One such region is the heterodimer formed between the first and second C-terminal domains of the endonuclease CPSF73, with those from the catalytically inactive CPSF100. Here we report the backbone and sidechain resonance assignments of a minimal C-terminal heterodimer of CPSF73–CPSF100 derived from the parasite <i>Encephalitozoon cuniculi</i>. The assignment process used several amino-acid specific labeling strategies, and the chemical shift values allow for secondary structure prediction.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"17 1","pages":"43 - 48"},"PeriodicalIF":0.9,"publicationDate":"2023-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-023-10118-6.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4021582","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-02-01DOI: 10.1007/s12104-022-10115-1
Glauce M. Barbosa, Maria A. Morando, Andrea T. Da Poian, Fabio C. L. Almeida
Dengue virus belongs to the Flaviviridae family, being responsible for an endemic arboviral disease in humans. It is an enveloped virus, whose genome is a positive-stranded RNA packaged by the capsid protein. Dengue virus capsid protein (DENVC) forms homodimers in solution organized in 4 α-helices and an intrinsically disordered N-terminal region. The N-terminal region is involved in the binding of membranous structures in host cells and in the recognition of nucleotides. Here we report the 1H, 15N and 13C resonance assignments of the DENVC with the deletion of the first 19 intrinsically disordered residues. The backbone chemical shift perturbations suggest changes in the α1 and α2 helices between full length and the truncated proteins.
{"title":"The 1H, 15N and 13C resonance assignments of dengue virus capsid protein with the deletion of the intrinsically disordered N-terminal region","authors":"Glauce M. Barbosa, Maria A. Morando, Andrea T. Da Poian, Fabio C. L. Almeida","doi":"10.1007/s12104-022-10115-1","DOIUrl":"10.1007/s12104-022-10115-1","url":null,"abstract":"<div><p>Dengue virus belongs to the <i>Flaviviridae</i> family, being responsible for an endemic arboviral disease in humans. It is an enveloped virus, whose genome is a positive-stranded RNA packaged by the capsid protein. Dengue virus capsid protein (DENVC) forms homodimers in solution organized in 4 α-helices and an intrinsically disordered N-terminal region. The N-terminal region is involved in the binding of membranous structures in host cells and in the recognition of nucleotides. Here we report the <sup>1</sup>H, <sup>15</sup>N and <sup>13</sup>C resonance assignments of the DENVC with the deletion of the first 19 intrinsically disordered residues. The backbone chemical shift perturbations suggest changes in the α1 and α2 helices between full length and the truncated proteins.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"17 1","pages":"23 - 26"},"PeriodicalIF":0.9,"publicationDate":"2023-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4022377","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-12-24DOI: 10.1007/s12104-022-10116-0
Justin A. Grennell, Kendra D. Jenkins, Kelvin B. Luther, John Glushka, Robert S. Haltiwanger, Megan A. Macnaughtan
NOTCH1 is a transmembrane receptor in metazoans that is linked to a variety of disorders. The receptor contains an extracellular domain (ECD) with 36 tandem epidermal growth factor-like (EGF) repeats. The ECD is responsible for intercellular signaling via protein–ligand interactions with neighboring cells. Each EGF repeat consists of approximately 40 amino acids and 3 conserved disulfide bonds. The Abruptex region (EGF24-29) is critical for NOTCH1 signaling and is known for its missense mutations. Certain EGF repeats are modified with the addition of O-linked glycans and many have calcium binding sites, which give each EGF repeat a unique function. It has been shown that the loss of the O-fucose site of EGF27 alters NOTCH1 activity. To investigate the role of glycosylation in the NOTCH1 signaling pathway, nuclear magnetic resonance spectroscopy has been employed to study the structures of EGF27 and its glycoforms. Here, we report the backbone and sidechain 1H, 15N, and 13C-resonance assignments of the unmodified EGF27 protein and the predicted secondary structure derived from the assigned chemical shifts.
{"title":"1H, 15N, 13C backbone and sidechain resonance assignments and secondary structure of mouse NOTCH1 EGF27","authors":"Justin A. Grennell, Kendra D. Jenkins, Kelvin B. Luther, John Glushka, Robert S. Haltiwanger, Megan A. Macnaughtan","doi":"10.1007/s12104-022-10116-0","DOIUrl":"10.1007/s12104-022-10116-0","url":null,"abstract":"<div><p>NOTCH1 is a transmembrane receptor in metazoans that is linked to a variety of disorders. The receptor contains an extracellular domain (ECD) with 36 tandem epidermal growth factor-like (EGF) repeats. The ECD is responsible for intercellular signaling via protein–ligand interactions with neighboring cells. Each EGF repeat consists of approximately 40 amino acids and 3 conserved disulfide bonds. The <i>Abruptex</i> region (EGF24-29) is critical for NOTCH1 signaling and is known for its missense mutations. Certain EGF repeats are modified with the addition of <i>O</i>-linked glycans and many have calcium binding sites, which give each EGF repeat a unique function. It has been shown that the loss of the <i>O</i>-fucose site of EGF27 alters NOTCH1 activity. To investigate the role of glycosylation in the NOTCH1 signaling pathway, nuclear magnetic resonance spectroscopy has been employed to study the structures of EGF27 and its glycoforms. Here, we report the backbone and sidechain <sup>1</sup>H, <sup>15</sup>N, and <sup>13</sup>C-resonance assignments of the unmodified EGF27 protein and the predicted secondary structure derived from the assigned chemical shifts.\u0000</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"17 1","pages":"27 - 35"},"PeriodicalIF":0.9,"publicationDate":"2022-12-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10116-0.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"5259852","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-12-21DOI: 10.1007/s12104-022-10117-z
Katherine M. Coburn, Braden Roth, Kristen M. Varney, France Carrier, David J. Weber
Heterogeneous ribonuclear protein A18 (hnRNP A18) is an RNA binding protein (RBP) involved in the hypoxic cellular stress response and regulation of cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) expression in melanoma, breast cancer, prostate cancer, and colon cancer solid tumors. hnRNP A18 is comprised of an N-terminal structured RNA recognition motif (RMM) and a C-terminal intrinsically disordered domain (IDD). Upon cellar stressors, such as UV and hypoxia, hnRNP A18 is phosphorylated by casein kinase 2 (CK2) and glycogen synthase kinase 3β (GSK-3β). After phosphorylation, hnRNP A18 translocates from the nucleus to the cytosol where it interacts with pro-survival mRNA transcripts for proteins such as hypoxia inducible factor 1α and CTLA-4. Both the hypoxic cellular response and modulation of immune checkpoints by cancer cells promote chemoradiation resistance and metastasis. In this study, the 1 H, 13 C, and 15 N backbone and sidechain resonances of the 172 amino acid hnRNP A18 were assigned sequence-specifically and provide a framework for future NMR-based drug discovery studies toward targeting hnRNP A18. These data will also enable the investigation of the dynamic structural changes within the IDD of hnRNP A18 upon phosphorylation by CK2 and GSK-3β to provide critical insight into the structure and function of IDDs.
{"title":"1H, 13C, and 15N assignments of the mRNA binding protein hnRNP A18","authors":"Katherine M. Coburn, Braden Roth, Kristen M. Varney, France Carrier, David J. Weber","doi":"10.1007/s12104-022-10117-z","DOIUrl":"10.1007/s12104-022-10117-z","url":null,"abstract":"<div><p>Heterogeneous ribonuclear protein A18 (hnRNP A18) is an RNA binding protein (RBP) involved in the hypoxic cellular stress response and regulation of cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) expression in melanoma, breast cancer, prostate cancer, and colon cancer solid tumors. hnRNP A18 is comprised of an N-terminal structured RNA recognition motif (RMM) and a C-terminal intrinsically disordered domain (IDD). Upon cellar stressors, such as UV and hypoxia, hnRNP A18 is phosphorylated by casein kinase 2 (CK2) and glycogen synthase kinase 3β (GSK-3β). After phosphorylation, hnRNP A18 translocates from the nucleus to the cytosol where it interacts with pro-survival mRNA transcripts for proteins such as hypoxia inducible factor 1α and CTLA-4. Both the hypoxic cellular response and modulation of immune checkpoints by cancer cells promote chemoradiation resistance and metastasis. In this study, the <sup>1</sup> H, <sup>13</sup> C, and <sup>15</sup> N backbone and sidechain resonances of the 172 amino acid hnRNP A18 were assigned sequence-specifically and provide a framework for future NMR-based drug discovery studies toward targeting hnRNP A18. These data will also enable the investigation of the dynamic structural changes within the IDD of hnRNP A18 upon phosphorylation by CK2 and GSK-3β to provide critical insight into the structure and function of IDDs.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"17 1","pages":"37 - 41"},"PeriodicalIF":0.9,"publicationDate":"2022-12-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10117-z.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"5116317","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-12-15DOI: 10.1007/s12104-022-10113-3
José Malanho Silva, Deborah Grifagni, Francesca Cantini, Mario Piccioli
CISD3 is a mitochondrial protein that contains two [2Fe–2S] clusters. This protein is overexpressed in some types of cancer, so it has emerged as a potential drug target. A detailed characterization of this protein is crucial to understand how CISD3 is involved in these physiopathologies. In this study, isotopically labeled human CISD3 was expressed in Escherichia coli. A set of double and triple resonance experiments performed with standard parameters/datasets provided the assignment of 40% of the HN resonances, 47% of Cα, and 46% of C′ resonances. Tailored paramagnetic HSQC, CON and CACO experiments extended up to 59% for HN, 70% for Cα and 69% for C′. The 1H, 13C and 15N NMR chemical shift assignment of human CISD3 is reported here.
{"title":"1H, 13C and 15N assignment of the human mitochondrial paramagnetic iron–sulfur protein CISD3","authors":"José Malanho Silva, Deborah Grifagni, Francesca Cantini, Mario Piccioli","doi":"10.1007/s12104-022-10113-3","DOIUrl":"10.1007/s12104-022-10113-3","url":null,"abstract":"<div><p>CISD3 is a mitochondrial protein that contains two [2Fe–2S] clusters. This protein is overexpressed in some types of cancer, so it has emerged as a potential drug target. A detailed characterization of this protein is crucial to understand how CISD3 is involved in these physiopathologies. In this study, isotopically labeled human CISD3 was expressed in <i>Escherichia coli</i>. A set of double and triple resonance experiments performed with standard parameters/datasets provided the assignment of 40% of the HN resonances, 47% of Cα, and 46% of C′ resonances. Tailored paramagnetic HSQC, CON and CACO experiments extended up to 59% for HN, 70% for Cα and 69% for C′. The <sup>1</sup>H, <sup>13</sup>C and <sup>15</sup>N NMR chemical shift assignment of human CISD3 is reported here.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"17 1","pages":"17 - 22"},"PeriodicalIF":0.9,"publicationDate":"2022-12-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10113-3.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4892434","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-10-29DOI: 10.1007/s12104-022-10112-4
Qinyan Song, Xiang-Qin Liu, Jan K. Rainey
The human MDM2 protein regulates the tumor suppressor protein p53 by restricting its transcriptional activity and by promoting p53 degradation. MDM2 is ubiquitously expressed, with its overexpression implicated in many forms of cancer. The inhibitory effects of MDM2 on p53 have been shown to involve its N-terminal p53-binding domain and its C-terminal RING domain. The presence of an intact central acidic domain of MDM2 has also been shown to regulate p53 ubiquitination, with this domain shown to directly interact with the p53 DNA-binding domain to regulate the DNA binding activity of p53. To date, little structural information has been obtained for the MDM2 acidic domain. Thus, to gain insight into the structure and function relationship of this region, we have applied solution-state NMR spectroscopy to characterize the segment of MDM2 spanning residues 215–300. These boundaries for the acidic domain were determined on the basis of consensus observed in multiple sequence alignment. Here, we report the 1H, 15N and 13C backbone and 13Cβ chemical shift assignments and steady-state {1H}-15N heteronuclear NOE enhancement factors as a function of residue for the acidic domain of MDM2. We show that this domain exhibits the hallmarks of being a disordered protein, on the basis both of assigned chemical shifts and residue-level backbone dynamics, with localized variation in secondary structure propensity inferred from chemical shift analysis.
{"title":"1H, 15N and 13C backbone resonance assignments of the acidic domain of the human MDM2 protein","authors":"Qinyan Song, Xiang-Qin Liu, Jan K. Rainey","doi":"10.1007/s12104-022-10112-4","DOIUrl":"10.1007/s12104-022-10112-4","url":null,"abstract":"<div><p>The human MDM2 protein regulates the tumor suppressor protein p53 by restricting its transcriptional activity and by promoting p53 degradation. MDM2 is ubiquitously expressed, with its overexpression implicated in many forms of cancer. The inhibitory effects of MDM2 on p53 have been shown to involve its N-terminal p53-binding domain and its C-terminal RING domain. The presence of an intact central acidic domain of MDM2 has also been shown to regulate p53 ubiquitination, with this domain shown to directly interact with the p53 DNA-binding domain to regulate the DNA binding activity of p53. To date, little structural information has been obtained for the MDM2 acidic domain. Thus, to gain insight into the structure and function relationship of this region, we have applied solution-state NMR spectroscopy to characterize the segment of MDM2 spanning residues 215–300. These boundaries for the acidic domain were determined on the basis of consensus observed in multiple sequence alignment. Here, we report the <sup>1</sup>H, <sup>15</sup>N and <sup>13</sup>C backbone and <sup>13</sup>C<sub>β</sub> chemical shift assignments and steady-state {<sup>1</sup>H}-<sup>15</sup>N heteronuclear NOE enhancement factors as a function of residue for the acidic domain of MDM2. We show that this domain exhibits the hallmarks of being a disordered protein, on the basis both of assigned chemical shifts and residue-level backbone dynamics, with localized variation in secondary structure propensity inferred from chemical shift analysis.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"17 1","pages":"9 - 16"},"PeriodicalIF":0.9,"publicationDate":"2022-10-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10112-4.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"5132648","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-10-22DOI: 10.1007/s12104-022-10111-5
Maria D. Politi, Angelo Gallo, Georgios Bouras, Maria Birkou, Bruno Canard, Bruno Coutard, Georgios A. Spyroulias
The genome of Hepatitis E virus (HEV) is 7.2 kilobases long and has three open reading frames. The largest one is ORF1, encoding a non-structural protein involved in the replication process, and whose processing is ill-defined. The ORF1 protein is a multi-modular protein which includes a macro domain (MD). MDs are evolutionarily conserved structures throughout all kingdoms of life. MDs participate in the recognition and removal of ADP-ribosylation, and specifically viral MDs have been identified as erasers of ADP-ribose moieties interpreting them as important players at escaping the early stages of host-immune response. A detailed structural analysis of the apo and bound to ADP-ribose state of the native HEV MD would provide the structural information to understand how HEV MD is implicated in virus-host interplay and how it interacts with its intracellular partner during viral replication. In the present study we present the high yield expression of the native macro domain of HEV and its analysis by solution NMR spectroscopy. The HEV MD is folded in solution and we present a nearly complete backbone and sidechains assignment for apo and bound states. In addition, a secondary structure prediction by TALOS + analysis was performed. The results indicated that HEV MD has a α/β/α topology very similar to that of most viral macro domains.
{"title":"1H, 13C, 15N backbone resonance assignment of apo and ADP-ribose bound forms of the macro domain of Hepatitis E virus through solution NMR spectroscopy","authors":"Maria D. Politi, Angelo Gallo, Georgios Bouras, Maria Birkou, Bruno Canard, Bruno Coutard, Georgios A. Spyroulias","doi":"10.1007/s12104-022-10111-5","DOIUrl":"10.1007/s12104-022-10111-5","url":null,"abstract":"<div><p>The genome of Hepatitis E virus (HEV) is 7.2 kilobases long and has three open reading frames. The largest one is ORF1, encoding a non-structural protein involved in the replication process, and whose processing is ill-defined. The ORF1 protein is a multi-modular protein which includes a macro domain (MD). MDs are evolutionarily conserved structures throughout all kingdoms of life. MDs participate in the recognition and removal of ADP-ribosylation, and specifically viral MDs have been identified as erasers of ADP-ribose moieties interpreting them as important players at escaping the early stages of host-immune response. A detailed structural analysis of the <i>apo</i> and bound to ADP-ribose state of the native HEV MD would provide the structural information to understand how HEV MD is implicated in virus-host interplay and how it interacts with its intracellular partner during viral replication. In the present study we present the high yield expression of the native macro domain of HEV and its analysis by solution NMR spectroscopy. The HEV MD is folded in solution and we present a nearly complete backbone and sidechains assignment for <i>apo</i> and bound states. In addition, a secondary structure prediction by TALOS + analysis was performed. The results indicated that HEV MD has a <i>α/β/α</i> topology very similar to that of most viral macro domains.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"17 1","pages":"1 - 8"},"PeriodicalIF":0.9,"publicationDate":"2022-10-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10111-5.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4880695","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-09-15DOI: 10.1007/s12104-022-10110-6
Nikolaos K. Fourkiotis, Periklis Charalampous, Aikaterini C. Tsika, Konstantina P. Kravvariti, Christos Sideras-Bisdekis, Angelo Gallo, Georgios A. Spyroulias
hPARP14 is a human ADP-ribosyl-transferase (ART) that belongs to the macroPARPs family, together with hPARP9 and hPARP15. It contains a tandem of three macro domains (MD) while each of them has different properties. The first one, namely MD1, has not been reported to exhibit a high binding affinity for ADP-ribose (ADPr) in contrast to the following two (MD2 and MD3). All three MDs exhibit an α/β/α sandwich-like fold as reported by the deposited crystallographic structures. MD2 and MD3 recognize mono-ADP-ribosylated (MARylated) but not poly-ADP-ribosylated (PARylated) substrates and thus they allow hPARP14 to bind its targets, which can be potentially MARylated by its catalytic domain (CD). hPARP14 participates in DNA damage repair process and immune response against viruses like SARS-CoV-2, which also harbors an MD fold. Furthermore, hPARP14 like the other two macroPARPs (hPARP9 and hPARP15), is implicated in numerous types of cancer, such as B-aggressive lymphoma and sarcoma, rendering its MDs as potential important drug targets. Herein, we report the complete NMR backbone and side chain assignment (1H, 13C, 15N) of hPARP14 MD2 in the free and ADPr bound states and the NMR chemical shift-based prediction of its secondary structure elements. This is the first reported NMR study of a hPARP macro domain, paving the way to screen by NMR chemical compounds which may alter the ability of hPARP14 to interact with its substrates affecting its function.
{"title":"NMR study of human macroPARPs domains: 1H, 15N and 13C resonance assignment of hPARP14 macro domain 2 in the free and the ADPr bound state","authors":"Nikolaos K. Fourkiotis, Periklis Charalampous, Aikaterini C. Tsika, Konstantina P. Kravvariti, Christos Sideras-Bisdekis, Angelo Gallo, Georgios A. Spyroulias","doi":"10.1007/s12104-022-10110-6","DOIUrl":"10.1007/s12104-022-10110-6","url":null,"abstract":"<div><p>hPARP14 is a human ADP-ribosyl-transferase (ART) that belongs to the macroPARPs family, together with hPARP9 and hPARP15. It contains a tandem of three macro domains (MD) while each of them has different properties. The first one, namely MD1, has not been reported to exhibit a high binding affinity for ADP-ribose (ADPr) in contrast to the following two (MD2 and MD3). All three MDs exhibit an α/β/α sandwich-like fold as reported by the deposited crystallographic structures. MD2 and MD3 recognize mono-ADP-ribosylated (MARylated) but not poly-ADP-ribosylated (PARylated) substrates and thus they allow hPARP14 to bind its targets, which can be potentially MARylated by its catalytic domain (CD). hPARP14 participates in DNA damage repair process and immune response against viruses like SARS-CoV-2, which also harbors an MD fold. Furthermore, hPARP14 like the other two macroPARPs (hPARP9 and hPARP15), is implicated in numerous types of cancer, such as B-aggressive lymphoma and sarcoma, rendering its MDs as potential important drug targets. Herein, we report the complete NMR backbone and side chain assignment (<sup>1</sup>H, <sup>13</sup>C, <sup>15</sup>N) of hPARP14 MD2 in the free and ADPr bound states and the NMR chemical shift-based prediction of its secondary structure elements. This is the first reported NMR study of a hPARP macro domain, paving the way to screen by NMR chemical compounds which may alter the ability of hPARP14 to interact with its substrates affecting its function.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"16 2","pages":"399 - 406"},"PeriodicalIF":0.9,"publicationDate":"2022-09-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10110-6.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4635275","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-09-12DOI: 10.1007/s12104-022-10105-3
Xiao Han, Maria Levkovets, Dmitry Lesovoy, Renhua Sun, Johan Wallerstein, Tatyana Sandalova, Tatiana Agback, Adnane Achour, Peter Agback, Vladislav Yu. Orekhov
Mucosa-associated lymphoid tissue protein 1 (MALT1) plays a key role in adaptive immune responses by modulating specific intracellular signalling pathways that control the development and proliferation of both T and B cells. Dysfunction of these pathways is coupled to the progress of highly aggressive lymphoma as well as to potential development of an array of different immune disorders. In contrast to other signalling mediators, MALT1 is not only activated through the formation of the CBM complex together with the proteins CARMA1 and Bcl10, but also by acting as a protease that cleaves multiple substrates to promote lymphocyte proliferation and survival via the NF-κB signalling pathway. Herein, we present the partial 1H, 13C Ile/Val/Leu-Methyl resonance assignment of the monomeric apo form of the paracaspase-IgL3 domain of human MALT1. Our results provide a solid ground for future elucidation of both the three-dimensional structure and the dynamics of MALT1, key for adequate development of inhibitors, and a thorough molecular understanding of its function(s).
粘膜相关淋巴组织蛋白1 (MALT1)通过调节控制T细胞和B细胞发育和增殖的特异性细胞内信号通路,在适应性免疫应答中发挥关键作用。这些途径的功能障碍与高度侵袭性淋巴瘤的进展以及一系列不同免疫疾病的潜在发展有关。与其他信号介质不同的是,MALT1不仅通过与CARMA1和Bcl10蛋白形成CBM复合物而被激活,而且还作为蛋白酶裂解多种底物,通过NF-κB信号通路促进淋巴细胞增殖和存活。在此,我们提出了部分1H, 13C Ile/Val/ leu -甲基共振分配的人MALT1的paracaspase-IgL3结构域的单体载子形式。我们的研究结果为未来阐明MALT1的三维结构和动力学提供了坚实的基础,这是充分开发抑制剂的关键,并对其功能进行彻底的分子理解。
{"title":"Assignment of IVL-Methyl side chain of the ligand-free monomeric human MALT1 paracaspase-IgL3 domain in solution","authors":"Xiao Han, Maria Levkovets, Dmitry Lesovoy, Renhua Sun, Johan Wallerstein, Tatyana Sandalova, Tatiana Agback, Adnane Achour, Peter Agback, Vladislav Yu. Orekhov","doi":"10.1007/s12104-022-10105-3","DOIUrl":"10.1007/s12104-022-10105-3","url":null,"abstract":"<div><p>Mucosa-associated lymphoid tissue protein 1 (MALT1) plays a key role in adaptive immune responses by modulating specific intracellular signalling pathways that control the development and proliferation of both T and B cells. Dysfunction of these pathways is coupled to the progress of highly aggressive lymphoma as well as to potential development of an array of different immune disorders. In contrast to other signalling mediators, MALT1 is not only activated through the formation of the CBM complex together with the proteins CARMA1 and Bcl10, but also by acting as a protease that cleaves multiple substrates to promote lymphocyte proliferation and survival via the NF-κB signalling pathway. Herein, we present the partial <sup>1</sup>H, <sup>13</sup>C Ile/Val/Leu-Methyl resonance assignment of the monomeric apo form of the paracaspase-IgL<sub>3</sub> domain of human MALT1. Our results provide a solid ground for future elucidation of both the three-dimensional structure and the dynamics of MALT1, key for adequate development of inhibitors, and a thorough molecular understanding of its function(s).</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"16 2","pages":"363 - 371"},"PeriodicalIF":0.9,"publicationDate":"2022-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10105-3.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4521118","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2022-09-09DOI: 10.1007/s12104-022-10109-z
Houman Ghasriani, Sara Ahmadi, Derek J. Hodgson, Yves Aubin
Monoclonal antibodies (mAbs) therapeutics are the largest and fastest growing class of biologic drugs, amongst which, the vast majority are immunoglobulin G1 (IgG1). Their antigen binding abilities are used for the treatment of immunologic diseases, cancer therapy, reversal of drug effects, and targeting viruses and bacteria. The high importance of therapeutic mAbs and their derivatives has called for the generation of well-characterized standards for method development and calibration. One such standard, the NISTmAb RM 8621 based on the antibody motavizumab, has been developed by the National Institute of Standards and Technologies (NIST) in the US. Here, we present the resonance assignment of the single chain variable fragment, NISTmAb-scFv, that was engineered by linking the variable domains of the heavy and light chains of the NISTmAb. Also, addition of a peptide, corresponding to the target antigen of motavizumab, to samples of NISTmAb-scFv has induced chemical shift perturbations on residues lining the antigen binding interface thereby indicating proper folding of the NISTmAb-scFv.
{"title":"Backbone and side-chain resonance assignments of the NISTmAb-scFv and antigen-binding study","authors":"Houman Ghasriani, Sara Ahmadi, Derek J. Hodgson, Yves Aubin","doi":"10.1007/s12104-022-10109-z","DOIUrl":"10.1007/s12104-022-10109-z","url":null,"abstract":"<div><p>Monoclonal antibodies (mAbs) therapeutics are the largest and fastest growing class of biologic drugs, amongst which, the vast majority are immunoglobulin G1 (IgG1). Their antigen binding abilities are used for the treatment of immunologic diseases, cancer therapy, reversal of drug effects, and targeting viruses and bacteria. The high importance of therapeutic mAbs and their derivatives has called for the generation of well-characterized standards for method development and calibration. One such standard, the NISTmAb RM 8621 based on the antibody motavizumab, has been developed by the National Institute of Standards and Technologies (NIST) in the US. Here, we present the resonance assignment of the single chain variable fragment, NISTmAb-scFv, that was engineered by linking the variable domains of the heavy and light chains of the NISTmAb. Also, addition of a peptide, corresponding to the target antigen of motavizumab, to samples of NISTmAb-scFv has induced chemical shift perturbations on residues lining the antigen binding interface thereby indicating proper folding of the NISTmAb-scFv.</p></div>","PeriodicalId":492,"journal":{"name":"Biomolecular NMR Assignments","volume":"16 2","pages":"391 - 398"},"PeriodicalIF":0.9,"publicationDate":"2022-09-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12104-022-10109-z.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4408120","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}