PGE1 and PGA1 bind to Nurr1 and activate its transcriptional function.

The orphan nuclear receptor Nurr1 is critical for the development, maintenance and protection of midbrain dopaminergic (mDA) neurons. Here we show that prostaglandin E1 (PGE1) and its dehydrated metabolite, PGA1, directly interact with the ligand-binding domain (LBD) of Nurr1 and stimulate its transcriptional function. We also report the crystallographic structure of Nurr1-LBD bound to PGA1 at 2.05 Å resolution. PGA1 couples covalently to Nurr1-LBD by forming a Michael adduct with Cys566, and induces notable conformational changes, including a 21° shift of the activation function-2 helix (H12) away from the protein core. Furthermore, PGE1/PGA1 exhibit neuroprotective effects in a Nurr1-dependent manner, prominently enhance expression of Nurr1 target genes in mDA neurons and improve motor deficits in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned mouse models of Parkinson's disease. Based on these results, we propose that PGE1/PGA1 represent native ligands of Nurr1 and can exert neuroprotective effects on mDA neurons, via activation of Nurr1's transcriptional function.

Special Issue "Selected Papers from the 8th Asia-Pacific NMR (APNMR) Symposium: Recent Advances in NMR Spectroscopy".

Asia-Pacific NMR (APNMR) has been an important scientific event in the region, engaging a large number of NMR scientists from academia and industries [...].

Self-association and conformational variation of NS5A domain 1 of hepatitis C virus.

Direct-acting antivirals (DAAs) targeting the non-structural 5A (NS5A) protein of the hepatitis C virus (HCV) are crucial drugs that have shown exceptional clinical success in patients. However, their mode of action (MoA) remains unclear, and drug-resistant HCV strains are rapidly emerging. It is critical to characterize the behaviour of the NS5A protein in solution, which can facilitate the development of new classes of inhibitors or improve the efficacy of the currently available DAAs. Using biophysical methods, including dynamic light scattering, size exclusion chromatography and chemical cross-linking experiments, we showed that the NS5A domain 1 from genotypes 1b and 1a of the HCV intrinsically self-associated and existed as a heterogeneous mixture in solution. Interestingly, the NS5A domain 1 from genotypes 1b and 1a exhibited different dynamic equilibria of monomers to higher-order structures. Using small-angle X-ray scattering, we studied the structural dynamics of the various states of the NS5A domain 1 in solution. We also tested the effect of daclatasvir (DCV), the most prominent DAA, on self-association of the wild and DCV-resistant mutant (Y93H) NS5A domain 1 proteins, and demonstrated that DCV induced the formation of large and irreversible protein aggregates that eventually precipitated out. This study highlights the conformational variability of the NS5A domain 1 of HCV, which may be an intrinsic structural behaviour of the HCV NS5A domain 1 in solution.

Molecular diversity and function of jasmintides from Jasminum sambac.

BACKGROUND: Jasmintides jS1 and jS2 from Jasminum sambac were previously identified as a novel family of cysteine-rich peptides (CRPs) with an unusual disulfide connectivity. However, very little else is known about jasmintides, particularly their molecular diversity and functions. Here, we report the discovery and characterization of a novel suite of jasmintides from J. sambac using transcriptomic, peptidomic, structural and functional tools. RESULTS: Transcriptomic analysis of leaves, flowers and roots revealed 14 unique jasmintide precursors, all of which possess a three-domain architecture comprising a signal peptide, a pro-domain and a mature jasmintide domain. Peptidomic analysis, using fractionated mixtures of jasmintides and chemical derivatization of cysteine to pseudolysine, trypsin digestion and MS/MS sequencing, revealed an additional 86 jasmintides, some of which were post-translationally modified. NMR analysis showed that jasmintide jS3 has three anti-parallel ß-strands with a three-disulfide connectivity of CysI-CysV, CysII-CysIV and CysIII-CysVI, which is similar to jasmintide jS1. Jasmintide jS3 was able to withstand thermal, acidic and enzymatic degradation and, importantly, exhibited antifeedant activity against mealworm Tenebrio molitor. CONCLUSION: Together, this study expands the existing library of jasmintides and furthers our understanding of the molecular diversity and cystine framework of CRPs as scaffolds and tools for engineering peptides targeting pests.

Structural basis of nucleic acid recognition by FK506-binding protein 25 (FKBP25), a nuclear immunophilin.

The nuclear immunophilin FKBP25 interacts with chromatin-related proteins and transcription factors and is suggested to interact with nucleic acids. Currently the structural basis of nucleic acid binding by FKBP25 is unknown. Here we determined the nuclear magnetic resonance (NMR) solution structure of full-length human FKBP25 and studied its interaction with DNA. The FKBP25 structure revealed that the N-terminal helix-loop-helix (HLH) domain and C-terminal FK506-binding domain (FKBD) interact with each other and that both of the domains are involved in DNA binding. The HLH domain forms major-groove interactions and the basic FKBD loop cooperates to form interactions with an adjacent minor-groove of DNA. The FKBP25-DNA complex model, supported by NMR and mutational studies, provides structural and mechanistic insights into the nuclear immunophilin-mediated nucleic acid recognition.

Nuclear receptor Nurr1 agonists enhance its dual functions and improve behavioral deficits in an animal model of Parkinson's disease.

Parkinson's disease (PD), primarily caused by selective degeneration of midbrain dopamine (mDA) neurons, is the most prevalent movement disorder, affecting 1-2% of the global population over the age of 65. Currently available pharmacological treatments are largely symptomatic and lose their efficacy over time with accompanying severe side effects such as dyskinesia. Thus, there is an unmet clinical need to develop mechanism-based and/or disease-modifying treatments. Based on the unique dual role of the nuclear orphan receptor Nurr1 for development and maintenance of mDA neurons and their protection from inflammation-induced death, we hypothesize that Nurr1 can be a molecular target for neuroprotective therapeutic development for PD. Here we show successful identification of Nurr1 agonists sharing an identical chemical scaffold, 4-amino-7-chloroquinoline, suggesting a critical structure-activity relationship. In particular, we found that two antimalarial drugs, amodiaquine and chloroquine stimulate the transcriptional function of Nurr1 through physical interaction with its ligand binding domain (LBD). Remarkably, these compounds were able to enhance the contrasting dual functions of Nurr1 by further increasing transcriptional activation of mDA-specific genes and further enhancing transrepression of neurotoxic proinflammatory gene expression in microglia. Importantly, these compounds significantly improved behavioral deficits in 6-hydroxydopamine lesioned rat model of PD without any detectable signs of dyskinesia-like behavior. These findings offer proof of principle that small molecules targeting the Nurr1 LBD can be used as a mechanism-based and neuroprotective strategy for PD.

Speckle reduction in quantitative phase imaging by generating spatially incoherent laser field at electroactive optical diffusers.

We studied quantitative phase imaging (QPI) using coherent laser illumination coupled with static and moving optical diffusers. The spatial coherence of a continuous-wave laser was controlled by tuning the particle size and the diffusion angle of optical diffusers for speckle-reduced 3D phase imaging of transparent objects. We used a common-path QPI configuration to investigate the coherent phase mapping of polystyrene micro-beads and breast cancer cells (MCF-7) under different degrees of coherent speckles. The proposed speckle reduction method could provide an avenue for enhancing lateral resolution and suppressing coherent artifacts of the phase images from QPI.

Studies on the Chitin Binding Property of Novel Cysteine-Rich Peptides from Alternanthera sessilis.

Hevein-like peptides make up a family of cysteine-rich peptides (CRPs) and play a role in plants in their defense against insects and fungal pathogens. In this study, we report the isolation and characterization of six hevein-like peptides, aSG1-G3 and aSR1-R3, collectively named altides from green and red varieties of Alternanthera sessilis, a perennial herb belonging to the Amaranthaceae family. Proteomic analysis of altides revealed they contain six cysteines (6C), seven glycines, four prolines, and a conserved chitin-binding domain (SXYGY/SXFGY). Thus far, only four 6C-hevein-like peptides have been isolated and characterized; hence, our study expands the existing library of these peptides. Nuclear magnetic resonance (NMR) study of altides showed its three disulfide bonds were arranged in a cystine knot motif. As a consequence of this disulfide arrangement, they are stable against thermal and enzymatic degradation. Gene cloning studies revealed altides contain a three-domain precursor with an endoplasmic reticulum signal peptide followed by a mature CRP domain and a short C-terminal tail. This indicates that the biosynthesis of altides is through the secretory pathway. (1)H NMR titration experiments showed that the 29-30-amino acid altides bind to chitin oligomers with dissociation constants in the micromolar range. Aromatic residues in the chitin-binding domain of altides were involved in the binding interaction. To the best of our knowledge, aSR1 is the smallest hevein-like peptide with a dissociation constant toward chitotriose comparable to those of hevein and other hevein-like peptides. Together, our study expands the existing library of 6C-hevein-like peptides and provides insights into their structure, biosynthesis, and interaction with chitin oligosaccharides.

Bh3 induced conformational changes in Bcl-Xl revealed by crystal structure and comparative analysis.

Apoptosis or programmed cell death is a regulatory process in cells in response to stimuli perturbing physiological conditions. The Bcl-2 family of proteins plays an important role in regulating homeostasis during apoptosis. In the process, the molecular interactions among the three members of this family, the pro-apoptotic, anti-apoptotic and BH3-only proteins at the mitochondrial outer membrane define the fate of a cell. Here, we report the crystal structures of the human anti-apoptotic protein Bcl-XL in complex with BH3-only BID(BH3) and BIM(BH3) peptides determined at 2.0 Å and 1.5 Å resolution, respectively. The BH3 peptides bind to the canonical hydrophobic pocket in Bcl-XL and adopt an alpha helical conformation in the bound form. Despite a similar structural fold, a comparison with other BH3 complexes revealed structural differences due to their sequence variations. In the Bcl-XL-BID(BH3) complex we observed a large pocket, in comparison with other BH3 complexes, lined by residues from helices α1, α2, α3, and α5 located adjacent to the canonical hydrophobic pocket. These results suggest that there are differences in the mode of interactions by the BH3 peptides that may translate into functional differences in apoptotic regulation.

RNAi-based therapeutic nanostrategy: IL-8 gene silencing in pancreatic cancer cells using gold nanorods delivery vehicles.

RNA interference (RNAi)-based gene silencing possesses great ability for therapeutic intervention in pancreatic cancer. Among various oncogene mutations, Interleukin-8 (IL-8) gene mutations are found to be overexpressed in many pancreatic cell lines. In this work, we demonstrate IL-8 gene silencing by employing an RNAi-based gene therapy approach and this is achieved by using gold nanorods (AuNRs) for efficient delivery of IL-8 small interfering RNA (siRNA) to the pancreatic cell lines of MiaPaCa-2 and Panc-1. Upon comparing to Panc-1 cells, we found that the dominant expression of the IL-8 gene in MiaPaCa-2 cells resulted in an aggressive behavior towards the processes of cell invasion and metastasis. We have hence investigated the suitability of using AuNRs as novel non-viral nanocarriers for the efficient uptake and delivery of IL-8 siRNA in realizing gene knockdown of both MiaPaCa-2 and Panc-1 cells. Flow cytometry and fluorescence imaging techniques have been applied to confirm transfection and release of IL-8 siRNA. The ratio of AuNRs and siRNA has been optimized and transfection efficiencies as high as 88.40 ± 2.14% have been achieved. Upon successful delivery of IL-8 siRNA into cancer cells, the effects of IL-8 gene knockdown are quantified in terms of gene expression, cell invasion, cell migration and cell apoptosis assays. Statistical comparative studies for both MiaPaCa-2 and Panc-1 cells are presented in this work. IL-8 gene silencing has been demonstrated with knockdown efficiencies of 81.02 ± 10.14% and 75.73 ± 6.41% in MiaPaCa-2 and Panc-1 cells, respectively. Our results are then compared with a commercial transfection reagent, Oligofectamine, serving as positive control. The gene knockdown results illustrate the potential role of AuNRs as non-viral gene delivery vehicles for RNAi-based targeted cancer therapy applications.

Cysteine-Rich Peptide Family with Unusual Disulfide Connectivity from Jasminum sambac.

Cysteine-rich peptides (CRPs) are natural products with privileged peptidyl structures that represent a potentially rich source of bioactive compounds. Here, the discovery and characterization of a novel plant CRP family, jasmintides from Jasminum sambac of the Oleaceae family, are described. Two 27-amino acid jasmintides (jS1 and jS2) were identified at the gene and protein levels. Disulfide bond mapping of jS1 by mass spectrometry and its confirmation by NMR spectroscopy revealed disulfide bond connectivity of C-1-C-5, C-2-C-4, and C-3-C-6, a cystine motif that has not been reported in plant CRPs. Structural determination showed that jS1 displays a well-defined structure framed by three short antiparallel ß-sheets. Genomic analysis showed that jasmintides share a three-domain precursor arrangement with a C-terminal mature domain preceded by a long pro-domain of 46 residues and an intron cleavage site between the signal sequence and pro-domain. The compact cysteine-rich structure together with an N-terminal pyroglutamic acid residue confers jasmintides high resistance to heat and enzymatic degradation, including exopeptidase treatment. Collectively, these results reveal a new plant CRP structure with an unusual cystine connectivity, which could be useful as a scaffold for designing peptide drugs.

Combination of pharmacophore hypothesis and molecular docking to identify novel inhibitors of HCV NS5B polymerase.

Hepatitis C virus (HCV) infection or HCV-related liver diseases are now shown to cause more than 350,000 deaths every year. Adaptability of HCV genome to vary its composition and the existence of multiple strains makes it more difficult to combat the emergence of drug-resistant HCV infections. Among the HCV polyprotein which has both the structural and non-structural regions, the non-structural protein NS5B RNA-dependent RNA polymerase (RdRP) mainly mediates the catalytic role of RNA replication in conjunction with its viral protein machinery as well as host chaperone proteins. Lack of such RNA-dependent RNA polymerase enzyme in host had made it an attractive and hotly pursued target for drug discovery efforts. Recent drug discovery efforts targeting HCV RdRP have seen success with FDA approval for sofosbuvir as a direct-acting antiviral against HCV infection. However, variations in drug-binding sites induce drug resistance, and therefore targeting allosteric sites could delay the emergence of drug resistance. In this study, we focussed on allosteric thumb site II of the non-structural protein NS5B RNA-dependent RNA polymerase and developed a five-feature pharmacophore hypothesis/model which estimated the experimental activity with a strong correlation of 0.971 & 0.944 for training and test sets, respectively. Further, the Güner-Henry score of 0.6 suggests that the model was able to discern the active and inactive compounds and enrich the true positives during a database search. In this study, database search and molecular docking results supported by experimental HCV viral replication inhibition assays suggested ligands with best fitness to the pharmacophore model dock to the key residues involved in thumbs site II, which inhibited the HCV 1b viral replication in sub-micro-molar range.

Solution structure of the cyclic-nucleotide binding homology domain of a KCNH channel.

The carboxy-terminal region of the KCNH family of potassium channels contains a cyclic-nucleotide binding homology domain (CNBHD) that is important for channel gating and trafficking. The solution structure of the CNBHD of the KCNH potassium of zebrafish was determined using solution NMR spectroscopy. This domain exists as a monomer under solution conditions and adopts a similar fold to that determined by X-ray crystallography. The CNBHD does not bind cAMP because residue Y740 blocks the entry of cyclic-nucleotide to the binding pocket. Relaxation results show that the CNBHD is rigid except that some residues in the loop between ß6 and ß7 are flexible. Our results will be useful to understand the gating mechanism of KCNH family members through the CNBHD.

Dual-site interactions of p53 protein transactivation domain with anti-apoptotic Bcl-2 family proteins reveal a highly convergent mechanism of divergent p53 pathways.

Molecular interactions between the tumor suppressor p53 and the anti-apoptotic Bcl-2 family proteins play an important role in the transcription-independent apoptosis of p53. The p53 transactivation domain (p53TAD) contains two conserved ΦXXΦΦ motifs (Φ indicates a bulky hydrophobic residue and X is any other residue) referred to as p53TAD1 (residues 15-29) and p53TAD2 (residues 39-57). We previously showed that p53TAD1 can act as a binding motif for anti-apoptotic Bcl-2 family proteins. In this study, we have identified p53TAD2 as a binding motif for anti-apoptotic Bcl-2 family proteins by using NMR spectroscopy, and we calculated the structures of Bcl-X(L)/Bcl-2 in complex with the p53TAD2 peptide. NMR chemical shift perturbation data showed that p53TAD2 peptide binds to diverse members of the anti-apoptotic Bcl-2 family independently of p53TAD1, and the binding between p53TAD2 and p53TAD1 to Bcl-X(L) is competitive. Refined structural models of the Bcl-X(L)·p53TAD2 and Bcl-2·p53TAD2 complexes showed that the binding sites occupied by p53TAD2 in Bcl-X(L) and Bcl-2 overlap well with those occupied by pro-apoptotic BH3 peptides. Taken together with the mutagenesis, isothermal titration calorimetry, and paramagnetic relaxation enhancement data, our structural comparisons provided the structural basis of p53TAD2-mediated interaction with the anti-apoptotic proteins, revealing that Bcl-X(L)/Bcl-2, MDM2, and cAMP-response element-binding protein-binding protein/p300 share highly similar modes of binding to the dual p53TAD motifs, p53TAD1 and p53TAD2. In conclusion, our results suggest that the dual-site interaction of p53TAD is a highly conserved mechanism underlying target protein binding in the transcription-dependent and transcription-independent apoptotic pathways of p53.

NMR solution structure of C2 domain of MFG-E8 and insights into its molecular recognition with phosphatidylserine.

MFG-E8 (also known as lactadherin), which is a secreted glycoprotein from a variety of cell types, possesses two EGF domains and tandem C domains with sequence homology to that of blood coagulation proteins factor V and factor VIII. MFG-E8 binds to phosphatidylserine (PS) in membranes with high affinity. We have recently shown that the C2 domain of MFG-E8 bears more specificity toward PS when compared with phosphatidylcholine (PC), another phospholipid thought to be involved in the immune function of phagocytes. In our current study, we have determined the solution structure of the C2 domain by nuclear magnetic resonance (NMR) spectroscopy, and characterized the molecular basis of binding between the C2 domain and PS by (31)P-NMR spectroscopy. Furthermore, we also verified that that positively charged and aromatic residues clustered in loops 1-3 of the C2 domain play key roles in recognizing PS in apoptotic cells.

Crystal structure of Plasmodium vivax FK506-binding protein 25 reveals conformational changes responsible for its noncanonical activity.

The malarial parasites currently remain one of the most dreadful parasites, which show increasing trend of drug resistance to the currently available antimalarial drugs. Thus, the need to identify and characterize new protein targets in these parasites can aid to design novel therapeutic strategies to combat malaria. Recently, the conserved FK506-binding protein family members with molecular weight of 35 kDa from Plasmodium falciparum and Plasmodium vivax (referred to as PfFKBP35 and PvFKBP35, respectively) were identified for drug targeting. Further data mining revealed a 25-kDa FKBP (FKBP25) family member present in the parasites. FKBP25 belongs to a unique class of FKBP, because it is a nuclear FKBP with multiple protein-binding partners. Apart from immune regulation, it is also known for its chaperoning role in various cellular processes such as transcription regulation and trafficking. Here, we present the biochemical characterization and 1.9-Å crystal structure of an N-terminal truncated FKBP25 from P. vivax (PvFKBP25(72-209)). The protein reveals the noncanonical nature with unique structural changes observed in the loops flanking the active site, concealing the binding pocket. Further, a potential calmodulin-binding domain, which is absent in human FKBP25, is observed in this protein. Although the functional implication of Plasmodium FKBP25 in malaria still remains elusive, we speculate that the notable conformational changes in its structure might serve as an overture in understanding its molecular mechanism.

Structural basis for the conserved binding mechanism of MDM2-inhibiting peptides and anti-apoptotic Bcl-2 family proteins.

The interaction between tumor suppressor p53 and the anti-apoptotic Bcl-2 family proteins serves a critical role in the transcription-independent apoptosis mechanism of p53. Our previous studies showed that an MDM2-inhibiting motif (residues 15-29) in the p53 transactivation domain (p53TAD) mediates the interaction with anti-apoptotic Bcl-2 family proteins. In this study, we provided structural models of the complexes between the MDM2-inhibiting p53TAD peptide and Mcl-1, Bcl-w, and Kaposi sarcoma-associated herpes virus (KSHV) Bcl-2 using NMR chemical shift perturbation data. The binding mode of the MDM2-inhibiting p53TAD peptide is highly conserved among the anti-apoptotic Bcl-2 family proteins despite their distinct specificities for pro-apoptotic Bcl-2 family proteins. We also identified the binding of a phage-display-derived MDM2-inhibiting peptide 12-1 to anti-apoptotic Bcl-XL protein by using NMR spectroscopy. The structural model of the Bcl-XL/12-1 peptide complex revealed that the conserved residues Phe4, Trp8, and Leu11 in the MDM2-inhibiting peptide fit into a hydrophobic cleft of Bcl-XL in a manner similar to that of pro-apoptotic Bcl-2 homology 3 (BH3) peptides. Our results shed light on the mechanism underlying dual-targeting of the FxxxWxxL-based α-helical motif to MDM2 and anti-apoptotic Bcl-2 family proteins for anticancer therapy.

Structural insights into substrate binding by PvFKBP35, a peptidylprolyl cis-trans isomerase from the human malarial parasite Plasmodium vivax.

The immunosuppressive drug FK506 binding proteins (FKBPs), an immunophilin family with the immunosuppressive drug FK506 binding property, exhibit peptidylprolyl cis-trans isomerase (PPIase) activity. While the cyclophilin-catalyzed peptidylprolyl isomerization of X-Pro peptide bonds has been extensively studied, the mechanism of the FKBP-mediated peptidylprolyl isomerization remains uncharacterized. Thus, to investigate the binding of FKBP with its substrate and the underlying catalytic mechanism of the FKBP-mediated proline isomerization, here we employed the FK506 binding domain (FKBD) of the human malarial parasite Plasmodium vivax FK506 binding protein 35 (PvFKBP35) and examined the details of the molecular interaction between the isomerase and a peptide substrate. The crystallographic structures of apo PvFKBD35 and its complex with the tetrapeptide substrate succinyl-Ala-Leu-Pro-Phe-p-nitroanilide (sALPFp) determined at 1.4 Å and 1.65 Å resolutions, respectively, showed that the substrate binds to PvFKBD35 in a cis conformation. Nuclear magnetic resonance (NMR) studies demonstrated the chemical shift perturbations of D55, H67, V73, and I74 residues upon the substrate binding. In addition, the X-ray crystal structure, along with the mutational studies, shows that Y100 is a key residue for the catalytic activity. Taken together, our results provide insights into the catalytic mechanism of PvFKBP35-mediated cis-trans isomerization of substrate and ultimately might aid designing substrate mimetic inhibitors targeting the malarial parasite FKBPs.

Obtusilactone B from Machilus Thunbergii targets barrier-to-autointegration factor to treat cancer.

Targeting specific molecules is a promising cancer treatment because certain types of cancer cells are dependent on specific oncogenes. This strategy led to the development of therapeutics that use monoclonal antibodies or small-molecule inhibitors. However, the continued development of novel molecular targeting inhibitors is required to target the various oncogenes associated with the diverse types and stages of cancer. Obtusilactone B is a butanolide derivative purified from Machilus thunbergii. In this study, we show that obtusilactone B functions as a small-molecule inhibitor that causes abnormal nuclear envelope dynamics and inhibits growth by suppressing vaccinia-related kinase 1 (VRK1)-mediated phosphorylation of barrier-to-autointegration factor (BAF). BAF is important in maintaining lamin integrity, which is closely associated with diseases that include cancer. Specific binding of obtusilactone B to BAF suppressed VRK1-mediated BAF phosphorylation and the subsequent dissociation of the nuclear envelope from DNA that allows cells to progress through the cell cycle. Obtusilactone B potently induced tumor cell death in vitro, indicating that specific targeting of BAF to block cell cycle progression can be an effective anticancer strategy. Our results demonstrate that targeting a major constituent of the nuclear envelope may be a novel and promising alternative approach to cancer treatment.

Macro histone H2A1.2 (macroH2A1) protein suppresses mitotic kinase VRK1 during interphase.

VRK1-mediated phosphorylation of histone H3 should be restricted in mitosis for consistent cell cycling, and defects in this process trigger cellular catastrophe. However, an interphasic regulator against VRK1 has not been actually investigated so far. Here, we show that the histone variant macrodomain-containing histone H2A1.2 functions as a suppressor against VRK1 during interphase. The level of macroH2A1.2 was markedly reduced in the mitotic phase, and the macroH2A1.2-mediated inhibition of histone H3 phosphorylation occurred mainly during interphase. We also found direct interaction and binding features between VRK1 and macroH2A1.2 by NMR spectroscopy. Hence, our findings might provide valuable insight into the underlying molecular mechanism regarding an epigenetic regulation of histone H3 during the cell cycle.