Envisaging the Structural Elevation in the Early Event of Oligomerization of Disordered Amyloid β Peptide

In Alzheimer’s disease (AD), amyloid β (Aβ) protein plays a detrimental role in neuronal injury and death. Recent in vitro and in vivo studies suggest that soluble oligomers of the Aβ peptide are neurotoxic. Structural properties of the oligomeric assembly, however, are largely unknown. Our present investigation established that the 40-residue-long Aβ peptide (Aβ40) became more helical, ordered, and compact in the oligomeric state, and both the helical and β-sheet components were found to increase significantly in the early event of oligomerization. The band-selective two-dimensional NMR analysis suggested that majority of the residues from sequence 12 to 22 gained a higher-ordered secondary structure in the oligomeric condition. The presence of a significant amount of helical conformation was confirmed by Raman bands at 1650 and 1336 cm^–1. Other residues remained mostly in the extended polyproline II (PPII) and less compact β-conformation space. In the event of maturation of the oligomers into an amyloid fiber, both the helical content and the PPII-like structural components declined and ∼72% residues attained a compact β-sheet structure. Interestingly, however, some residues remained in the collagen triple helix/extended 2.5₁-helix conformation as evidenced by the amide III Raman signature band at 1272 cm^–1. Molecular dynamics analysis using an optimized potential for liquid simulation force field with the peptide monomer indicated that some of the residues may have preferences for helical conformation and this possibly contributed in the event of oligomer formation, which eventually became a β-sheet-rich amyloid fiber

Spin-Noise-Detected Two-Dimensional Fourier-Transform NMR Spectroscopy

We introduce two-dimensional NMR spectroscopy detected by recording and processing the noise originating from nuclei that have not been subjected to any radio frequency excitation. The method relies on cross-correlation of two noise blocks that bracket the evolution and mixing periods. While the sensitivity of the experiment is low in conventional NMR setups, spin-noise-detected NMR spectroscopy has great potential for use with extremely small numbers of spins, thereby opening a way to nanoscale multidimensional NMR spectroscopy

Liaison between Myristoylation and Cryptic EF-Hand Motif Confers Ca²⁺ Sensitivity to Neuronal Calcium Sensor‑1

Many members of the neuronal calcium sensor (NCS) protein family have a striking coexistence of two characteristics, that is, N-myristoylation and the cryptic EF-1 motif. We investigated the rationale behind this correlation in neuronal calcium sensor-1 (NCS-1) by restoring Ca²⁺ binding ability of the disabled EF-1 loop by appropriate mutations. The concurrence of canonical EF-1 and N-myristoylation considerably decreased the overall Ca²⁺ affinity, conformational flexibility, and functional activation of downstream effecter molecules (i.e., PI4Kβ). Of a particular note, Ca²⁺ induced conformational change (which is the first premise for a CaBP to be considered as sensor) is considerably reduced in myristoylated proteins in which Ca²⁺-binding to EF-1 is restored. Moreover, Ca²⁺, which otherwise augments the enzymatic activity of PI4Kβ (modulated by NCS-1), leads to a further decline in the modulated PI4Kβ activity by myristoylated mutants (with canonical EF-1) pointing toward a loss of Ca²⁺ signaling and specificity at the structural as well as functional levels. This study establishes the presence of the strong liaison between myristoylation and cryptic EF-1 in NCS-1. Breaking this liaison results in the failure of Ca²⁺ specific signal transduction to downstream effecter molecules despite Ca²⁺ binding. Thus, the EF-1 disability is a prerequisite in order to append myristoylation signaling while preserving structural robustness and Ca²⁺ sensitivity/specificity in NCS-1

Aggregation-Prone Near-Native Intermediate Formation during Unfolding of a Structurally Similar Nonlenticular βγ-Crystallin Domain

The folding and unfolding of structurally similar proteins belonging to a family have long been a focus of investigation of the structure–(un)­folding relationship. Such studies are yet to reach a consensus about whether structurally similar domains follow common or different unfolding pathways. Members of the βγ-crystallin superfamily, which consists of structurally similar proteins with limited sequence similarity from diverse life forms spanning microbes to mammals, form an appropriate model system for exploring this relationship further. We selected a new member, Crybg3_D3, the third βγ-crystallin domain of non-lens vertebrate protein Crybg3 from mouse brain. The crystal structure determined at 1.86 Å demonstrates that the βγ-crystallin domain of Crybg3 resembles more closely the lens βγ-crystallins than the microbial crystallins do. However, interestingly, this structural cousin follows a quite distinct (un)­folding pathway via formation of an intermediate state. The intermediate species is in a nativelike conformation with variation in flexibility and tends to form insoluble aggregates. The individual domains of lens βγ-crystallins (and microbial homologues) do not follow such an unfolding pattern. Thus, even the closest members of a subfamily within a superfamily do not necessarily follow similar unfolding paths, suggesting the divergence acquired by these domains, which could be observed only by unfolding. Additionally, this study provides insights into the modifications that this domain has undergone during its recruitment into the non-lens tissues in vertebrates