Mechanisms Of Protein Misfolding Research Articles

Massively parallel molecular dynamics simulations of lysozyme unfolding

We have performed molecular dynamics simulations for a total duration of more than 10 µs (with most molecular trajectories being 1 µs in duration) to study the effect of a single mutation on hen lysozyme protein stability and denaturing, using an IBM Blue Gene/L™ supercomputer. One goal of this study was to assess the use of certain force fields to reproduce experimental results of protein unfolding using thermal denaturing techniques. A second and more important goal was to gain microscopic insights into the mechanism of protein misfolding using both thermal and chemical denaturing techniques. We found that the thermal denaturing results were robust and reproducible with various force fields. The chemical denaturing results explained why the single amino-acid mutation on residue Trp62 causes the disruption of long-range interactions in the tertiary structure. Simulation results revealed that the Trp62 residue was the key to a cooperative long-range interaction within the wild-type protein. Specifically, Trp62 acts as a bridge between two neighboring basic residues through a π-type H-bond or π-cation interaction to form an Arg-Trp-Arg “sandwich-like” structure. Our findings support the general conclusions of the experiment and provide an interesting molecular depiction of the disruption of the long-range interactions.

Tyrosine side chains as an electrochemical probe of stacked β-sheet protein conformations

The in vivo formation of β-pleated protein aggregates underlies a number of fatal neurodegenerative disorders, such as Alzheimer disease. Since molecular mechanisms of protein misfolding and aggregation remain poorly understood, this has been calling for many diverse biophysical tools capable of addressing different dynamic and conformational aspects of the phenomenon. The two model polypeptides used in this study are poly( l-tyrosine) and insulin. According to FT-IR spectra, poly( l-tyrosine) produced two distinct types of films with dominant either disordered or antiparallel β-sheet conformations depending on carrier solvent used for film's deposition. Electrochemical analysis of both the types of polypeptide films by the means of cyclic voltammetry and differential pulse voltammetry proved that different electrochemical behaviour of the tyrosine residues is determined by the conformation of polypeptide chains. We have rationalized this difference in terms of varying electrochemical accessibility of Tyr residues in each structure. We have also carried out spectral and electrochemical characterization of insulin β-sheet-rich amyloid fibrils. It appears that the detectable electrochemical response of the protein stems from the presence of four tyrosine residues per insulin monomer. Since hydrophobic residues, among them tyrosines play an important role in the formation of protein amyloid fibrils, but, on a molecular level, may be also critical in explaining neurotoxic properties of aggregates, their electrochemical properties may become a very valuable complementary tool in biophysical studies on protein misfolding.

Expansion of amino acid homo-sequences in proteins: insights into the role of amino acid homo-polymers and of the protein context in aggregation.

Expansion of amino acid homo-sequences, such as polyglutamines or polyalanines, in proteins has been directly implicated in various degenerative diseases through a mechanism of protein misfolding and aggregation. However, it is still unclear how the nature of the expansion and the protein context influence the tendency of a protein to aggregate. Here, we have addressed these questions using spinocerebellar ataxia type-3 (ATX3) protein, the best characterised of the polyglutamine proteins, chosen as a model system. Using a transfected mammalian cell line, we demonstrate that ATX3 aggregation is noticeably reduced by deletion or replacement of regions other than the polyglutamine tract. The nature of the amino acid homo-sequences also has a strong influence on aggregation. From our studies, we draw general conclusions on the effect of the protein architecture and of the amino acid homo-sequence on pathology.

Recent Developments in the Understanding and Treatment of Neurodegenerative Disorders Involving Protein Conformational Misfolding and Amyloid Formation

Conformational disease represents an intriguing but devastating class of neurodegenerative disorders that includes prion disease, Alzheimers disease, Parkinsons disease and Huntingtons disease. Although symptoms, on-set times and prognosis among the diseases can vary markedly, the deposition of neurotoxic protein aggregates is a significant commonality, and as such is an attractive therapeutic target. Understanding the mechanisms of protein misfolding and deposition in these conditions is critical to developing effective diagnostic and therapeutic agents. This review serves as an update for the sister publication “Protein Conformational Misfolding and Amyloid Formation: Characteristics of a New Class of Disorders that Include Alzheimers and Prion Diseases” in Curr. Med. Chem. 2002, 9, 1751-62, and focuses primarily on recent developments in understanding prion disease and Alzheimers disease in context with other conformational disease. New research in amyloid-related therapeutic strategies is also discussed.

New insights into the mechanisms of protein misfolding and aggregation in amyloidogenic diseases derived from pressure studies.

Hydrostatic pressure is a robust tool for studying the thermodynamics of protein folding and protein interactions, as well as the dynamics and structure of folding intermediates. One of the main innovations obtained from using high pressure is the stabilization of folding intermediates such as molten-globule conformations, thus providing a unique opportunity for characterizing their structure and dynamics. Equally important is the prospect of understanding protein misfolding diseases by using pressure to populate partially folded intermediates at the junction between productive and off-pathway folding, which may give rise to misfolded proteins, aggregates, and amyloids. High hydrostatic pressure (HHP) has also been used to dissociate nonamyloid aggregates and inclusion bodies. In many proteins, the competition between correct folding and misfolding can lead to formation of insoluble aggregates, an important problem for the biotechnology industry and for human pathologies such as amyloidosis, Alzheimer's, Parkinson's, prion's, and tumor diseases. The diversity of diseases that result from protein misfolding has made this theme an important research focus for pharmaceutical and biotechnology companies. The use of high-pressure promises to contribute to the identification of the mechanisms behind these defects and creation of therapies against these diseases.

Unfolding the role of protein misfolding in neurodegenerative diseases.

Recent evidence indicates that diverse neurodegenerative diseases might have a common cause and pathological mechanism — the misfolding, aggregation and accumulation of proteins in the brain, resulting in neuronal apoptosis. Studies from different disciplines strongly support this hypothesis and indicate that a common therapy for these devastating disorders might be possible. The aim of this article is to review the literature on the molecular mechanism of protein misfolding and aggregation, its role in neurodegeneration and the potential targets for therapeutic intervention in neurodegenerative diseases. Many questions still need to be answered and future research in this field will result in exciting new discoveries that might impact other areas of biology.

Many naturally occurring mutations of myelin proteolipid protein impair its intracellular transport.

The primary structure of the proteolipid protein (PLP) from the central nervous system (CNS) myelin of mammals is highly conserved with only three amino acid differences between the mouse, rat, dog, bovine, and human proteins. Furthermore, within a particular species no polymorphisms in the protein have been identified. Recent interest has focused on the targeting of PLP in oligodendrocytes and the role that mutant forms of this protein play in generating dysmyelinating or hypomyelinating diseases. We previously expressed the human cDNA encoding PLP in transiently transfected Cos-7 cells and characterized the subcellular distribution of the protein in this simple heterologous system. In the current study we have used the same paradigm to examine the effect of five missense mutations in the PLP gene on processing of the encoded protein. The mutations chosen span the carboxy-terminal half of PLP and encompass that part of the protein in which most mutations have been identified. Our results show that transport of all mutations examined was arrested in the secretory pathway at an early stage, causing the mutant proteins to accumulate in the endoplasmic reticulum. Thus, a common mechanism of protein misfolding and failure of PLP to reach the cell surface of oligodendrocytes rather than the inability of the mutant protein to perform some crucial function at the cell surface may be responsible for the diseases caused by many PLP mutations. Our results, together with those of others, prompt us to speculate that the pathobiology observed in PLP mutants may result from oligodendrocyte cell death caused by the accumulation of misfolded protein in the endoplasmic reticulum. This speculation is consistent with the observations that oligodendrocytes bearing misfolded PLP, as in the jimpy mutant, proliferate but die rapidly while oligodendrocytes from PLP deletion survive and produce a myelin-like membrane which lacks PLP.