Disorder in CENP-ACse4 tail-chaperone interaction facilitates binding with Ame1/Okp1 at the kinetochore.

The centromere is epigenetically marked by a histone H3 variant-CENP-A. The budding yeast CENP-A called Cse4, consists of an unusually long N-terminus that is known to be involved in kinetochore assembly. Its disordered chaperone, Scm3 is responsible for the centromeric deposition of Cse4 as well as in the maintenance of a segregation-competent kinetochore. In this study, we show that the Cse4 N-terminus is intrinsically disordered and interacts with Scm3 at multiple sites, and the complex does not gain any substantial structure. Additionally, the complex forms a synergistic association with an essential inner kinetochore component (Ctf19-Mcm21-Okp1-Ame1), and a model has been suggested to this effect. Thus, our study provides mechanistic insights into the Cse4 N-terminus-chaperone interaction and also illustrates how intrinsically disordered proteins mediate assembly of complex multiprotein networks, in general.

Disorder in the Human Skp1 Structure is the Key to its Adaptability to Bind Many Different Proteins in the SCF Complex Assembly.

Skp1(S-phase kinase-associated protein 1 - Homo sapiens) is an adapter protein of the SCF(Skp1-Cullin1-Fbox) complex, which links the constant components (Cul1-RBX) and the variable receptor (F-box proteins) in Ubiquitin E3 ligase. It is intriguing how Skp1 can recognise and bind to a variety of structurally different F-box proteins. For practical reasons, previous efforts have used truncated Skp1, and thus it has not been possible to track the crucial aspects of the substrate recognition process. In this background, we report the solution structure of the full-length Skp1 protein determined by NMR spectroscopy for the first time and investigate the sequence-dependent dynamics in the protein. The solution structure reveals that Skp1 has an architecture: ß1-ß2-H1-H2-L1-H3-L2-H4-H5-H6-H7(partially formed) and a long tail-like disordered C-terminus. Structural analysis using DALI (Distance Matrix Alignment) reveals conserved domain structure across species for Skp1. Backbone dynamics investigated using NMR relaxation suggest substantial variation in the motional timescales along the length of the protein. The loops and the C-terminal residues are highly flexible, and the (R2/R1) data suggests µs-ms timescale motions in the helices as well. Further, the dependence of amide proton chemical shift on temperature and curved profiles of their residuals indicate that the residues undergo transitions between native state and excited state. The curved profiles for several residues across the length of the protein suggest that there are native-like low-lying excited states, particularly for several C-terminal residues. Our results provide a rationale for how the protein can adapt itself, bind, and get functionally associated with other proteins in the SCF complex by utilising its flexibility and conformational sub-states.

Catching Nucleosome by Its Decorated Tails Determines Its Functional States.

The fundamental packaging unit of chromatin, i.e., nucleosome, consists of ∼147 bp of DNA wrapped around a histone octamer composed of the core histones, H2A, H2B, H3, and H4, in two copies each. DNA packaged in nucleosomes must be accessible to various machineries, including replication, transcription, and DNA damage repair, implicating the dynamic nature of chromatin even in its compact state. As the tails protrude out of the nucleosome, they are easily accessible to various chromatin-modifying machineries and undergo post-translational modifications (PTMs), thus playing a critical role in epigenetic regulation. PTMs can regulate chromatin states via charge modulation on histones, affecting interaction with various chromatin-associated proteins (CAPs) and DNA. With technological advancement, the list of PTMs is ever-growing along with their writers, readers, and erasers, expanding the complexity of an already intricate epigenetic field. In this review, we discuss how some of the specific PTMs on flexible histone tails affect the nucleosomal structure and regulate the accessibility of chromatin from a mechanistic standpoint and provide structural insights into some newly identified PTM-reader interaction.

Co-Administration of Aluminium Hydroxide Nanoparticles and Protective Antigen Domain 4 Encapsulated Non-Ionic Surfactant Vesicles Show Enhanced Immune Response and Superior Protection against Anthrax.

Aluminium salts have been the adjuvant of choice in more than 100 licensed vaccines. Here, we have studied the synergistic effect of aluminium hydroxide nanoparticles (AH np) and non-ionic surfactant-based vesicles (NISV) in modulating the immune response against protective antigen domain 4 (D4) of Bacillus anthracis. NISV was prepared from Span 60 and cholesterol, while AH np was prepared from aluminium chloride and sodium hydroxide. AH np was co-administered with NISV encapsulating D4 (NISV-D4) to formulate AHnp/NISV-D4. The antigen-specific immune response of AHnp/NISV-D4 was compared with that of commercial alhydrogel (alhy) co-administered with NISV-D4 (alhydrogel/NISV-D4), NISV-D4, AHnp/D4, and alhydrogel/D4. Co-administration of NISV-D4 with AH np greatly improved the D4-specific antibody titer as compared to the control groups. Based on IgG isotyping and ex vivo cytokine analysis, AHnp/NISV-D4 generated a balanced Th1/Th2 response. Furthermore, AH np/NISV-D4 showed superior protection against anthrax spore challenge in comparison to other groups. Thus, we demonstrate the possibility of developing a novel combinatorial nanoformulation capable of augmenting both humoral and cellular response, paving the way for adjuvant research.