Peptide-Based Strategies: Combating Alzheimer's Amyloid β Aggregation through Ergonomic Design and Fibril Disruption.

Amyloidosis of amyloid-β (Aβ) triggers a cascade of events, leading to oxidative damage and neuronal death. Therefore, inhibiting Aβ amyloidosis or disrupting the matured fibrils is the primary target to combat progressive Alzheimer's disease (AD) pathogenesis. Here, we undertake optimization strategies to improve the antiamyloid efficiency of our previously reported NF11 (NAVRWSLMRPF) peptide.

Enhancing amyloid beta inhibition and disintegration by natural compounds: A study utilizing spectroscopy, microscopy and cell biology.

Amyloid proteins and peptides play a pivotal role in the etiology of various neurodegenerative diseases, including Alzheimer's disease (AD). Synthetically designed small molecules/ peptides/ peptidomimetics show promise towards inhibition of various kinds of amyloidosis. However, exploration of compounds isolated from natural extracts having such potential is lacking.

14-3-3 interaction with phosphodiesterase 8A sustains PKA signaling and downregulates the MAPK pathway.

The cAMP/PKA and mitogen-activated protein kinase (MAPK) signaling cascade control many cellular processes and are highly regulated for optimal cellular responses upon external stimuli. Phosphodiesterase 8A (PDE8A) is an important regulator that inhibits signaling via cAMP-dependent PKA by hydrolyzing intracellular cAMP pool. Conversely, PDE8A activates the MAPK pathway by protecting CRAF/Raf1 kinase from PKA-mediated inhibitory phosphorylation at Ser259 residue, a binding site of scaffold protein 14-3-3.

Coomassie brilliant blue G-250 acts as a potential chemical chaperone to stabilize therapeutic insulin.

Our studies show Coomassie Brilliant Blue G-250 as a promising chemical chaperone that stabilises the α-helical native human insulin conformers, disrupting their aggregation. Furthermore, it also increases the insulin secretion. This multipolar effect coupled with its non-toxic nature could be useful for developing highly bioactive, targeted and biostable therapeutic insulin.

Naringenin-Functionalized Gold Nanoparticles and Their Role in α-Synuclein Stabilization.

Misfolding and self-assembly of several intrinsically disordered proteins into ordered β-sheet-rich amyloid aggregates emerged as hallmarks of several neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. Here we show how the naringenin-embedded nanostructure effectively retards aggregation and fibril formation of α-synuclein, which is strongly associated with the pathology of Parkinson's-like diseases. Naringenin is a polyphenolic compound from a plant source, and in our current investigation, we reported the one-pot synthesis of naringenin-coated spherical and monophasic gold nanoparticles (NAR-AuNPs) under optimized conditions.

Modulatory role of copper on hIAPP aggregation and toxicity in presence of insulin.

Aggregation of the human islets amyloid polypeptide, or hIAPP, is linked to β-cell death in type II diabetes mellitus (T2DM). Different pancreatic β-cell environmental variables such as pH, insulin and metal ions play a key role in controlling the hIAPP aggregation. Since insulin and hIAPP are co-secreted, it is known from numerous studies that insulin suppresses hIAPP fibrillation by preventing the initial dimerization process.

Mechanistic insight into functionally different human islet polypeptide (hIAPP) amyloid: the intrinsic role of the C-terminal structural motifs.

Targeting amyloidosis requires high-resolution insight into the underlying mechanisms of amyloid aggregation. The sequence-specific intrinsic properties of a peptide or protein largely govern the amyloidogenic propensity. Thus, it is essential to delineate the structural motifs that define the subsequent downstream amyloidogenic cascade of events.

Solvent Relaxation NMR: A Tool for Real-Time Monitoring Water Dynamics in Protein Aggregation Landscape.

Solvent dynamics strongly induce the fibrillation of an amyloidogenic system. Probing the solvation mechanism is crucial as it enables us to predict different proteins' functionalities, such as the aggregation propensity, structural flexibility, and toxicity. This work shows that a straightforward NMR method in conjunction with phenomenological models gives a global and qualitative picture of water dynamics at different concentrations and temperatures.

Effect of PEGylation on Host Defense Peptide Complexation with Bacterial Lipopolysaccharide.

Conjugation with poly(ethylene glycol) ("PEGylation") is a widely used approach for improving the therapeutic propensities of peptide and protein drugs through prolonging bloodstream circulation, reducing toxicity and immunogenicity, and improving proteolytic stability. In the present study, we investigate how PEGylation affects the interaction of host defense peptides (HDPs) with bacterial lipopolysaccharide (LPS) as well as HDP suppression of LPS-induced cell activation. In particular, we investigate the effects of PEGylation site for KYE28 (KYEITTIHNLFRKLTHRLFRRNFGYTLR), a peptide displaying potent anti-inflammatory effects, primarily provided by its N-terminal part.

Praja1 ubiquitin ligase facilitates degradation of polyglutamine proteins and suppresses polyglutamine-mediated toxicity.

A network of chaperones and ubiquitin ligases sustain intracellular proteostasis and is integral in preventing aggregation of misfolded proteins associated with various neurodegenerative diseases. Using cell-based studies of polyglutamine (polyQ) diseases, spinocerebellar ataxia type 3 (SCA3) and Huntington's disease (HD), we aimed to identify crucial ubiquitin ligases that protect against polyQ aggregation. We report here that Praja1 (PJA1), a Ring-H2 ubiquitin ligase abundantly expressed in the brain, is diminished when polyQ repeat proteins (ataxin-3/huntingtin) are expressed in cells.

Comparison of Synthetic Neuronal Model Membrane Mimics in Amyloid Aggregation at Atomic Resolution.

Alzheimer's disease (AD) is a severe neurodegenerative disorder caused by abnormal accumulation of toxic amyloid plaques of the amyloid-beta (Aβ) or the tau proteins in the brain. The plaque deposition leading to the collapse of the cellular integrity is responsible for a myriad of surface phenomena acting at the neuronal lipid interface. Recent years have witnessed dysfunction of the blood-brain barriers (BBB) associated with AD.

Targeted inhibition of amyloidogenesis using a non-toxic, serum stable strategically designed cyclic peptide with therapeutic implications.

Amyloidogenic disorders are currently rising as a global health issue, prompting more and more studies dedicated to the development of effective targeted therapeutics. The innate affinity of these amyloidogenic proteins towards the biomembranes adds further complexities to the systems. Our previous studies have shown that biologically active peptides can effectively target amyloidogenesis serving as an efficient therapeutic alternative in several amyloidogenic disorders.

Self-Assembly and Neurotoxicity of β-Amyloid (21-40) Peptide Fragment: The Regulatory Role of GxxxG Motifs.

The three GxxxG repeating motifs from the C-terminal region of β-amyloid (Aβ) peptide play a significant role in regulating the aggregation kinetics of the peptide. Mutation of these glycine residues to leucine greatly accelerates the fibrillation process but generates a varied toxicity profile. Using an array of biophysical techniques, we demonstrated the uniqueness of the composite glycine residues in these structural repeats.

Lipopolysaccharide from Gut Microbiota Modulates α-Synuclein Aggregation and Alters Its Biological Function.

Altered intestinal permeability has been correlated with Parkinson's pathophysiology in the enteric nervous system, before manifestations in the central nervous system (CNS). The inflammatory endotoxin or lipopolysaccharide (LPS) released by gut bacteria is known to modulate α-synuclein amyloidogenesis through the formation of intermediate nucleating species. Here, biophysical techniques in conjunction with microscopic images revealed the molecular interaction between lipopolysaccharide and α-synuclein that induce rapid nucleation events.

Bipartite Role of Heat Shock Protein 90 (Hsp90) Keeps CRAF Kinase Poised for Activation.

CRAF kinase maintains cell viability, growth, and proliferation by participating in the MAPK pathway. Unlike BRAF, CRAF requires continuous chaperoning by Hsp90 to retain MAPK signaling. However, the reason behind the continuous association of Hsp90 with CRAF is still elusive.

Inhibition of Insulin Amyloid Fibrillation by a Novel Amphipathic Heptapeptide: MECHANISTIC DETAILS STUDIED BY SPECTROSCOPY IN COMBINATION WITH MICROSCOPY.

The aggregation of insulin into amyloid fibers has been a limiting factor in the development of fast acting insulin analogues, creating a demand for excipients that limit aggregation. Despite the potential demand, inhibitors specifically targeting insulin have been few in number. Here we report a non-toxic and serum stable-designed heptapeptide, KR7 (KPWWPRR-NH), that differs significantly from the primarily hydrophobic sequences that have been previously used to interfere with insulin amyloid fibrillation.

Biophysical Characterization of Essential Phosphorylation at the Flexible C-Terminal Region of C-Raf with 14-3-3ζ Protein.

Phosphorylation at the C-terminal flexible region of the C-Raf protein plays an important role in regulating its biological activity. Auto-phosphorylation at serine 621 (S621) in this region maintains C-Raf stability and activity. This phosphorylation mediates the interaction between C-Raf and scaffold protein 14-3-3ζ to activate the downstream MEK kinase pathway.

Hsp70 clears misfolded kinases that partitioned into distinct quality-control compartments.

Hsp70 aids in protein folding and directs misfolded proteins to the cellular degradation machinery. We describe discrete roles of Hsp70,SSA1 as an important quality-control machinery that switches functions to ameliorate the cellular environment. SSA1 facilitates folding/maturation of newly synthesized protein kinases by aiding their phosphorylation process and also stimulates ubiquitylation and degradation of kinases in regular protein turnover or during stress when kinases are denatured or improperly folded.

Role of molecular chaperones in biogenesis of the protein kinome.

Molecular chaperones promote polypeptide folding in cells by protecting newly made and otherwise misfolded proteins against aggregation or degradation by the ubiquitin proteasome pathway. The roles of Saccharomyces cerevisiae Cdc37 and Ydj1 molecular chaperones are described in this chapter. We focus on biogenesis of protein kinases that require several different molecular chaperones for their proper folding.

Ubr1 and Ubr2 function in a quality control pathway for degradation of unfolded cytosolic proteins.

Quality control systems facilitate polypeptide folding and degradation to maintain protein homeostasis. Molecular chaperones promote folding, whereas the ubiquitin/proteasome system mediates degradation. We show here that Saccharomyces cerevisiae Ubr1 and Ubr2 ubiquitin ligases promote degradation of unfolded or misfolded cytosolic polypeptides.

Hsp110 chaperones control client fate determination in the hsp70-Hsp90 chaperone system.

Heat shock protein 70 (Hsp70) plays a central role in protein homeostasis and quality control in conjunction with other chaperone machines, including Hsp90. The Hsp110 chaperone Sse1 promotes Hsp90 activity in yeast, and functions as a nucleotide exchange factor (NEF) for cytosolic Hsp70, but the precise roles Sse1 plays in client maturation through the Hsp70-Hsp90 chaperone system are not fully understood. We find that upon pharmacological inhibition of Hsp90, a model protein kinase, Ste11DeltaN, is rapidly degraded, whereas heterologously expressed glucocorticoid receptor (GR) remains stable.

Ydj1 protects nascent protein kinases from degradation and controls the rate of their maturation.

Ydj1 is a Saccharomyces cerevisiae Hsp40 molecular chaperone that functions with Hsp70 to promote polypeptide folding. We identified Ydj1 as being important for maintaining steady-state levels of protein kinases after screening several chaperones and cochaperones in gene deletion mutant strains. Pulse-chase analyses revealed that a portion of Tpk2 kinase was degraded shortly after synthesis in a ydj1Delta mutant, while the remainder was capable of maturing but with reduced kinetics compared to the wild type.

Uncoupling of hormone-dependence from chaperone-dependence in the L701H mutation of the androgen receptor.

The mechanisms underlying androgen receptor (AR)-mediated progression of prostate cancer following androgen ablation have yet to be fully determined. On this basis we screened naturally occurring mutants of human AR for hormone-independent activity using a yeast model system. An initial screen of 43 different mutants revealed that ARs having a Leu701His mutation (AR(L701H)) exhibited hormone-independent activation of a lacZ reporter gene.

Cdc37 has distinct roles in protein kinase quality control that protect nascent chains from degradation and promote posttranslational maturation.

Cdc37 is a molecular chaperone that functions with Hsp90 to promote protein kinase folding. Analysis of 65 Saccharomyces cerevisiae protein kinases ( approximately 50% of the kinome) in a cdc37 mutant strain showed that 51 had decreased abundance compared with levels in the wild-type strain. Several lipid kinases also accumulated in reduced amounts in the cdc37 mutant strain.

Molecular chaperones and protein kinase quality control.

The Hsp90-Cdc37 chaperone pair has special responsibility for folding of protein kinases. This function has made Hsp90 a target for new chemotherapeutic approaches, and several compounds are currently being tested for their ability to inhibit many different kinases simultaneously. Not all kinases are sensitive to these inhibitors, however, and this difference might depend on how each kinase interacts with Hsp90 and Cdc37 during folding of the nascent chain and thereafter.