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Abstract: Protein misfolding and amyloid formations are associated with many neurodegenerative and systemic diseases. The discovery of Alzheimer’s disease and its association with the accumulation of Amyloid-β (Aβ) peptides in the plaques uncovered the pleiotropic nature of peptides/ proteins. As of today, more than 50 proteins/ peptides are reported to form amyloids or amyloid-like protein aggregates under different conditions, establishing that amyloid formation could be a generic property of many proteins. In principle, under certain conditions, all the proteins have this property to form amyloid-like aggregates, which can be toxic or non-toxic. The extensive research in this direction led to an understanding of the ubiquitous nature of amyloids. Mounting evidences suggest that processed foods, particularly protein-rich foods, could be a plethora of amyloids or amyloid-like protein aggregates. Many are reported to be toxic, and their consumption raises health concerns. The assimilation of dietary proteins in the human body largely depends upon their conformational states and the digestive integrity of the gastrointestinal system. Amyloids or amyloid-like protein aggregates are usually protease resistant, and their presence in foods is likely to reduce nutritional value. Several biochemical and biophysical factors, commonly evident in various food processing industries, such as high temperature, the addition of acid, etc., are likely to induce the formation of protease-resistant protein aggregates. Aging significantly alters gastrointestinal health, predisposing aged individuals to be more susceptible to protein aggregation-related diseases. Consumption of foods containing such protein aggregates will lead to a poor supply of essential amino acids and might exaggerate the amyloid-related disease etiology. On the other hand, the gut microbiome plays a crucial role during pathological events leading to the development of Alzheimer’s and Parkinson’s diseases. The activity of gastrointestinal proteases, pH change, gut microbiome, and intestinal epithelium integrity would largely determine the outcome of consuming foods loaded with such protein aggregates. The current review outlines the recent development in this area and a new perspective for designing safe protein-rich diets for healthy nutrition.

Infectious diseases are one of the leading causes of human deaths worldwide. They have devastated human life for a very long time; however, with advancements in diagnostics, prevention, and therapeutic approaches, they were controlled to a manageable extent during the second half of the 20th century. It was assumed that humankind has successfully defeated the threat of infectious diseases. However, many infectious diseases have undergone re-emergence and are now a major cause of concern. Besides, there is the emergence of several new infectious diseases. These diseases are termed re-emerging and emerging infectious diseases, and collectively account for more than 20 infectious diseases. World Health Organization has identified these diseases as the predominant health hazard faced by human beings. Owing to this situation, there is an urgent need to revisit infectious diseases and make efforts towards the development of anti-microbial drugs for emerging and reemerging microbial diseases. The present book chapter aims to provide a comprehensive account of re-emerging and newly emerging infectious diseases and the paradigm shift of antimicrobial drugs in the 21st century. It is expected to provide useful insight into this important research and development area.

The loss of crystallins solubility with aging and the formation of amyloid-like aggregates is considered the hallmark characteristic of cataract pathology. The present study was carried out to assess the effect of temperature on the soluble lens protein and the formation of protein aggregates with typical amyloid characteristics. The soluble fraction of lens proteins was subjected for heat treatment in the range of 40–60 °C, and the nature of protein aggregates was assessed by using Congo red (CR), thioflavin T (ThT), and 8-anilinonaphthalene-1-sulfonic acid (ANS) binding assays, circular dichroism (CD), Fourier-transform infrared (FT-IR) spectroscopy, and transmission electron microscopy (TEM). The heat-treated protein samples displayed a substantial bathochromic shift (≈15 nm) in the CR's absorption maximum (λmax) and increased ThT and ANS binding. The heat treatment of lens soluble proteins results in the formation of nontoxic, β-sheet rich, non-fibrillar, protein aggregates similar to the structures evident in the insoluble fraction of proteins isolated from the cataractous lens. The data obtained from the present study suggest that the exposure of soluble lens proteins to elevated temperature leads to the formation of non-fibrillar aggregates, establishing the role of amyloid in the heat-induced augmentation of cataracts pathology.

Purified soya bean proteins (glycinin and conglycinin) are known to form amyloid-like aggregates in vitro at a higher temperature. Soya beans (chunks) are textured proteinaceous vegetables made from defatted soya flour by heating it above 100°C and extruding under high pressure. Therefore, it was assumed that subjecting the soya bean proteins to high temperatures raises the possibility of forming amyloids or amyloid-like protein aggregates. Hence, the present study aimed to examine the presence of amyloid-like protein aggregates in soya beans. The isolated protein aggregates from hydrated soya beans displayed typical characteristics of amyloids, such as the red shift in the absorption maximum (λ

The age-dependent loss of solubility and aggregation of crystallins constitute the pathological hallmarks of cataract. Several biochemical and biophysical factors are responsible for the reduction of crystallins’ solubility and formation of irreversible protein aggregates, which display amyloid-like characteristics. The present study reports the heat-induced aggregation of soluble proteins isolated from human cataract lenses and the formation of amyloid-like structures. Exposure of protein at 55 °C for 4 h resulted in extensive (≈ 60%) protein aggregation. The heat-induced protein aggregates displayed substantial (≈ 20 nm) redshift in the wavelength of maximum absorption (λmax) of Congo red (CR) and increase in Thioflavin T (ThT) fluorescence emission intensity, indicating the presence of amyloid-like structures in the heat-induced protein aggregates. Subsequently, the addition of trehalose resulted in substantial inhibition of heat-induced aggregation and the formation of amyloid-like structure. The ability of trehalose to inhibit the heat-induced aggregation was found to be linearly dependent upon its concentration used. The optimum effect was observed in the presence of 30–40% (w/v) trehalose where the aggregated was found to be reduced from 60 to 30%. The present study demonstrated the ability to trehalose to inhibit the protein aggregation and interfere with the formation of amyloid-like structures.

Peptides constitute an essential component of all organisms' protein homeostasis ranging from bacteria, plants, and animals. They have organically been evolved to perform a wide range of essential functions, including their role as neurotransmitters, antimicrobial peptides (AMPs), and hormones. AMPs are short peptides synthesized by almost all organisms, implicated in guarding the host from various microbial infections. Their inherent ability to differentiate the target microbes from the host confers them excellent prospects in fighting against microbial infections and affirming their robust therapeutic potential against numerous drug-resistant microbes. Amyloidogenic peptides (AMYs) represent another class of short peptides armed with inherent aggregation propensity and form fibrillar aggregates rich in cross β-sheet structure. They are often involved in various degenerative diseases, including Alzheimer's disease (AD), Parkinson's disease (PD), and type-2 diabetes. Although these two distinct classes of peptides (i.e., AMPs and AMYs) appear to be functionally divergent, recent studies suggest that they possess a significant degree of structural and functional reciprocity. Consistent with this, many AMPs display amphiphilic nature, and hence, they can facilitate membrane remodeling processes, such as pore formation and fusion, similar to AMYs. The mounting evidence suggests the inherent ability of AMPs to self-assemble to form amyloid-like structures. On the other hand, the demonstration of antimicrobial properties of AMYs in their monomeric conformation provides a hint about the existence of an evolutionary linkage between these two classes of peptides. The congregation of specific amino acids to form aggregation-prone regions in a protein/peptide might have served as an evolutionary reservoir from which AMPs and AMYs were consecutively evolved. The current article reviews the fundamental features of the AMPs, AMYs, and their inter-relatedness and emerging paradigm for their inter-conversion.

Cottage cheese, extensively consumed worldwide, contains coagulated milk protein (casein), produced through boiling and acidification of milk. Casein forms amyloid or amyloid-like structures at high temperatures and low pH. Due to the similarities in the preparation of casein amyloids and cottage cheese, we hypothesized the presence of amyloid or amyloid-like protein aggregates in cottage cheese. To examine this hypothesis, cottage cheese was prepared from cow (Bos indicus) milk and isolated amyloids through a water extraction method. The isolated protein aggregates displayed typical characteristics of amyloids, such as a bathochromic shift in the wavelength of maximum absorption (λmax) of Congo red (CR), high thioflavin T (ThT) binding, increased surface hydrophobicity, and high β-sheet structure. However, they did not show antibacterial activity and toxic properties against erythrocytes. Our study revealed that the heat-treatment and subsequent acidification during cottage cheese preparation lead to the formation of non-toxic amyloid-like aggregates.

Alzheimer’s disease (AD) is regarded as one of the hallmark outcomes of an aging brain, and it is characterized by sequential loss of memory and changes in brain mass. In the advanced stage of the disease, a patient hardly recognizes the family members and fellow beings. Alzheimer’s disease represents one of the major contributors to the development of dementia, which is characterized by the formation of amyloid plaques and neurofibrillary tangles in brain tissues. In the recent past, several efforts pertaining to drug discovery have been made to interfere with the development and progression of Alzheimer’s disease. However, so far, no FDA approved drug is available to prevent or cure the disease. In recent days, generous attention is being given to formulating food-based therapeutic strategies to interfere with disease development and mitigate the symptoms. These food-based therapeutic formulations are loaded with critical bioactive compounds that provide specific health benefits in addition to supplementing fundamental nutrition. In the future, an amalgamation of modern knowledge of bioactive molecules with the traditional wisdom of food formulation will lead to the designing of a disease-specific therapeutic regime to prevent and cure numerous chronic diseases. Ayurveda, a herbal-based system of traditional medicines and therapeutic strategies, native to India and the Indian subcontinent, offers a number of unique strategies to prevent, delay, and cure various mental illnesses, including AD. It epitomizes the holistic method of treatment and promotes equilibrium in various dimensions of human existence and unification of the body, mind, and soul. There are several formulations that have claimed to slow down the age-related mental illnesses, and in general, enhance memory retention and improve cognitive behaviors. Some of the key bioactive molecules that serve as potential ingredients of nutraceuticals are obtained from native plants such as Turmeric (Curcuma longa), Tea (Camilla sinensis), Gingko (Ginkgo biloba), Cinnamon (Cinnamomum zeylanicum), Black pepper (Piper nigrum L.), Ginger (Zingiber officinale), etc. The efficiency and efficacy of these plants, which have been tested for the last 2000 years, show the richness of the ancient and ethnic Indian traditional knowledge. However, the effects of these preparations on biochemical processes of disease development at the molecular level are yet to be worked out. Nevertheless, in recent years, the needs for detailed pharmacological and toxicological studies have received the attention of researchers across the globe, and investigations are going on to substantiate the claimed pharmacological and therapeutic effects. This chapter will provide outlines of the current understanding of molecular mechanisms and hypotheses leading to the development of age-related mental illnesses and dementia in particular. It will also explore the existing therapeutic strategies, their limitations, and exploration of new possibilities, including the application of nutraceuticals, which in addition to providing basic nutrition signal, would also provide neuroprotective health benefits with minimum side effects.

Alzheimer's disease (AD) and cataract represent two common protein misfolding diseases closely associated with aging. Growing evidence suggests that these two diseases may be interrelated with each other through cross-sequence interactions between β-amyloid (Aβ) peptide and the short aggregating peptides derived from proteolytic breakdown of α-crystallin. αΑ(66–80) is one of several peptides produced by the proteolytic breakdown of α-crystallin in aged eye lens. Although it is evident that the Aβ(1–40) and αΑ(66–80) coexist in aged eye lenses and both the peptides are known to form macromolecular assemblies, their cross-sequence interaction and the seeding behavior are not known. In this study, the aggregation behavior of αΑ(66–80) has been examined in the presence of Aβ(1–40) on using thioflavin T (ThT) based aggregation kinetics. The presence of monomeric Aβ(1–40) augmented the aggregation kinetics of αΑ(66–80) and reduced the lag time of αΑ(66–80) aggregation. However, the addition of Aβ(1–40) or αΑ(66–80) fibrils (seeds) didn't result in any change in the rate of αΑ(66–80) aggregation. In this in vitro study, we could show that the presence Aβ(1–40) has substantial effect on the aggregation of αΑ(66–80), which suggests a possible interaction between AD and cataract pathologies.

The emergence of novel coronavirus SARS-CoV-2 is responsible for causing coronavirus disease-19 (COVID-19) imposing serious threat to global public health. Infection of SARS-CoV-2 to the host cell is characterized by direct translation of positive single stranded (+ ss) RNA to form large polyprotein polymerase 1ab (pp1ab), which acts as precursor for a number of nonstructural and structural proteins that play vital roles in replication of viral genome and biosynthesis of new virus particles. The maintenance of viral protein homeostasis is essential for continuation of viral life cycle in the host cell. To test whether the protein homeostasis of SARS-CoV-2 can be disrupted by inducing specific protein aggregation, we made an effort to examine whether the viral proteome contains any aggregation prone regions (APRs) that can be explored for inducing toxic protein aggregation specifically in viral proteins and without affecting the host cell. This curiosity leads to the identification of several (> 70) potential APRs in SARS-CoV-2 proteome. The length of the APRs ranges from 5 to 25 amino acid residues. Nearly 70% of total APRs investigated are relatively smaller and found to be in the range of 5-10 amino acids. The maximum number of ARPs (> 50) was observed in pp1ab. On the other hand, the structural proteins such as, spike (S), nucleoprotein (N), membrane (M) and envelope (E) proteins also possess APRs in their primary structures which altogether constitute 30% of the total APRs identified. Our findings may provide new windows of opportunities to design specific peptide-based, anti-SARS-CoV-2 therapeutic molecules against COVID-19.

Biofilms have significant role in microbial persistence, antibiotic resistance and chronic infections; consequently, there is a pressing need for development of novel “anti-biofilm strategies”. One of the fundamental mechanisms involved in biofilm formation is protein-protein interactions of ‘amyloid like proteins’ (ALPs) in extracellular matrix. Such interactions could be potential targets for development of novel anti-biofilm strategies; therefore, assessing the structural features of these interactions could be of great scientific value. Characterization of biomolecular interaction with conventional structure biology tools including X-Ray diffraction and Nuclear Magnetic Resonance is technically challenging, expensive and time-consuming. In contrast, modelling such interactions is time-efficient, economical and might provide deeper understanding of structural basis of interactions. Therefore, during the present study, protein-protein interaction of TasA(28-261)–TapA(33-253) (which is a decisive process for biofilm formation by Bacillus subtilis) was modeled using in silico approaches viz., molecular modelling, protein-protein docking and molecular dynamics simulations. Results identified amino-acid residues present within intrinsically disordered regions of both proteins to be critical for interaction. These results were further supported with PCA and FEL analyses. Results presented here represent novel finding and we hypothesize that aa identified during the present study could be targeted for inhibition of biofilm formation by B. subtilis.

Biofilms have a significant role in microbial persistence, antibiotic resistance, and chronic infections; consequently, there is a pressing need for development of novel "anti-biofilm strategies." One of the fundamental mechanisms involved in biofilm formation is protein-protein interactions of "amyloid-like proteins" (ALPs) in the extracellular matrix. Such interactions could be potential targets for development of novel anti-biofilm strategies; therefore, assessing the structural features of these interactions could be of great scientific value. Characterization of structural features the of protein-protein interaction with conventional structure biology tools including X-ray diffraction and nuclear magnetic resonance is technically challenging, expensive, and time-consuming. In contrast, modeling such interactions is time-efficient and economical, and might provide deeper understanding of structural basis of interactions. Although it is often acknowledged that molecular modeling methods have varying accuracy, their careful implementation with supplementary verification methods can provide valuable insight and directions for future studies. With this reasoning, during the present study, the protein-protein interaction of TasA

The aggregation of β-crystallins in the human eye lens constitutes a critical step during the development of cataract. We anticipated that the presence of Aggregation-Prone Regions (APRs) in their primary structure, which might be responsible for conformational change required for the self-assembly. To examine the presence of APRs, we systematically analyzed the primary structures of β-crystallins. Out of seven subtypes, the βB1-crystallin found to possess the highest aggregation score with 9 APRs in its primary structure. To confirm the amyloidogenic nature of these newly identified APRs, we further studied the aggregation behavior of one of the APRs spanning from 174 to 180 residues (174LWVYGFS180) of βB1-crystallin, which is referred as βB1(174-180). Under in vitro conditions, the synthetic analogue of βB1(174-180) peptide formed visible aggregates and displayed high Congo red (CR) bathochromic shift, Thioflavin T (ThT) binding and fibrilar morphology under transmission electron microscopy, which are the typical characteristics of amyloids. Further, the aggregated βB1(174-180) was found to induce aggregation of the soluble fraction of proteins isolated from the human cataractous lens. This observation suggests that the presence of APRs in βB1-crystallin might be serving as one of the intrinsic supplementary factors responsible for constitutive aggregation behavior of βB1-crystallin and development of cataract.

Pheromone peptides are an important component of bacterial quorum-sensing system. The pheromone peptide cOB1 (VAVLVLGA) of native commensal Enterococcus faecalis has also been identified as an antimicrobial peptide (AMP) and reported to kill the prototype clinical isolate strain of E. faecalis V583. In this study, the pheromone peptide cOB1 has shown to form amyloid-like structures, a characteristic which is never reported for a pheromone peptide so far. With in silico analysis, the peptide was predicted to be highly amyloidogenic. Further, under experimental conditions, cOB1 formed aggregates displaying characteristics of amyloid structures such as bathochromic shift in Congo red absorbance, enhancement in thioflavin T fluorescence, and fibrillar morphology under transmission electron microscopy. This novel property of pheromone peptide cOB1 may have some direct effects on the binding of the pheromone to the receptor cells and subsequent conjugative transfer, making this observation more important for the therapeutics, dealing with the generation of virulent and multidrug-resistant pathogenic strains.

Protegrin-4 (PG-4) is a member of the porcine leukocyte protegrins family of cysteine-rich antimicrobial peptides (AMPs) isolated from Sus scrofa. It consists of 18 amino acid residues and works as a part of innate immune system. In this study, we examined the intrinsic aggregation propensity of this AMP using multiple computational algorithms, namely, TANGO, AGGRESCAN, FOLDAMYLOID, AMYLPRED, and ZYGGREGATOR, and found that the peptide is predicted to have a high propensity for the β sheet formation that disposes this peptide to be amyloidogenic. Under in vitro conditions, PG-4 formed visible aggregates and displayed the hallmark properties of typical amyloids such as enhanced binding of Congo red, increased fluorescence with Thioflavin-T, and fibrillar morphology under transmission electron microscopy. Then we examined its antimicrobial activity against Bacillus subtilis and found that the aggregated peptide retained its antimicrobial activity. Additionally, the aggregates remain non-toxic to the HEK293 and Caco2 cells. Our study suggests that the inherent aggregation properties of AMP can rationally be explored as a potential source of peptide-based antimicrobials with enhanced stability.

N-terminal verkurzte, Pyroglutamat-modifizierte Varianten des Aβ-Peptids sind naturlich vorkommende chemische Modifikationen bei der Alzheimerschen Krankheit. Wir zeigen hier, dass die beiden Modifikationen die Lange der Fibrillen und den Ubergangsmittelpunkt der thermischen Entfaltung der Fibrillen signifikant herabsetzen. Die Peptidkonformation in den Fibrillen bleibt jedoch weitestgehend unbeeinflusst. Diese Beobachtung lasst darauf schliesen, dass der unmodifizierte N-Terminus die Fibrillen vor mechanischem Stress und Fragmentierung schutzt. Sie erklart ebenfalls die hohe Wahrscheinlichkeit von Pyroglutamat-modifiziertem Peptid, kleinere, besonders toxische Aggregate zu bilden.

Cn-AMP2 is an antimicrobial peptide derived from liquid endosperm of coconut (Cocos nucifera). It consists of 11 amino acid residues and predicted to have high propensity for β-sheet formation that disposes this peptide to be amyloidogenic. In the present study, we have examined the amyloidogenic propensities of Cn-AMP2 in silico and then tested the predictions under in vitro conditions. The in silico study revealed that the peptide possesses high amyloidogenic propensity comparable with Aβ. Upon solubilisation and agitation in aqueous buffer, Cn-AMP2 forms visible aggregates that display bathochromic shift in the Congo red absorbance spectra, strong increase in thioflavin T fluorescence and fibrillar morphology under transmission electron microscopy. All these properties are typical of an amyloid fibril derived from various proteins/peptides including Aβ.

Extravillous trophoblast (EVT) migration and invasion is the crucial step for normal placental development. IL-11 is a cytokine regulating cell migration and invasion in cells and is a critical factor for successful implantation of an embryo. Higher expression of thrombin receptor PAR-1 was reported in early pregnancy. The precise role of thrombin in trophoblast functions is not well understood. In this study, we asked whether thrombin can induce IL-11 secretion in trophoblasts if yes, which physiological cell functions are possibly affected? In this study, HTR-8/SVneo cells, which were originally derived from first-trimester villous explants of early pregnancy were used as the extravillous trophoblast (EVT) model. BeWo cells were used as the cytotrophoblast model. For gene silencing, qPCR and ELISA, each experiment was performed in triplicates for minimum three times. Here, we found that thrombin stimulates IL-11 gene expression and protein secretion in HTR-8/SVneo cells but not in BeWo cells. PAR-1 was the only receptor which was highly expressed in HTR-8/SVneo cells. Thrombin-mediated expression and secretion of IL-11 were mainly activated via PAR-1 receptor. Rac1, but not Rho-kinase activation is required for thrombin-induced IL-11 secretion. We also found that thrombin stimulation significantly enhanced cell migration that was inhibited after silencing the IL-11 gene. In conclusion, this study demonstrates the role of thrombin in regulating human EVT migration via IL-11 secretion. We propose that thrombin might regulate EVT migration through the decidua and spiral artery remodeling. Failure of thrombin-dependent EVT migration results in pregnancy disorder, such as preeclampsia.

Amyloid fibrils are linear polypeptide aggregates with a cross-β structure. These fibrils are best known for their association with neurodegenerative diseases, such as Alzheimer's or Parkinson's, but they may also be used by living organisms as functional units, e.g. in the synthesis of melanin or in the formation of bacterial biofilms. About a decade ago, in a search for semen factors that modulate infection by HIV-1 (a sexually transmitted virus and the causative agent of the acquired immune deficiency syndrome (AIDS)), it was demonstrated that semen harbors amyloid fibrils capable of markedly increasing HIV infection rates. This discovery not only created novel opportunities to prevent sexual HIV-1 transmission but also stimulated research to unravel the natural role of these factors. We discuss here the identification of these intriguing structures, their molecular properties, and their effects on both sexually transmitted diseases and reproductive health. Moreover, we review strategies to antagonize semen amyloid to prevent sexual transmission of viruses.

Human semen contains a large number of macromolecules, including proteins/enzymes and carbohydrates, regulating and protecting sperm cells. Proteomic analysis of human seminal fluid led to the discovery of semen amyloids derived from short peptide fragments of the proteins prostatic acid phosphatase (PAP) and semenogelin (SG) which are known to play a crucial role in enhancing HIV infection. However, the relevance of their existence in human semen and role in maintaining sperm behavior remains unclear. Distinct physiological, biochemical, and biophysical attributes might cause these amyloids to influence sperm behavior positively or negatively, affecting fertilization or other reproductive processes. We assessed the direct effect of amyloids derived from a PAP248–286 fragment, on sperm motility and viability, which are crucial parameters for assessment of sperm quality in semen. Co-incubation of human sperm with PAP248–286 amyloids at normal physiological concentrations formed in buffer led to significant reduction in sperm viability, though approximately a 10× higher concentration was needed to show a similar effect with amyloid formed in seminal fluid. Both forms of PAP248–286 amyloid also had a significant impact on sperm motility at physiological levels, in agreement with a previous report. Our study suggests that PAP248–286 amyloids can directly influence sperm motility and viability in a concentration-dependent manner. We hypothesise that the direct toxic effect of PAP248–286 amyloid is normally mitigated by other seminal fluid ingredients, but that in pathological conditions, where PAP248–286 concentrations are elevated and it plays a role in determining sperm health and viability, with relevance for male fertility as well as sterility.

Crystallin is a major protein present in eye lens. Peptide fragment αA(66-80) derived from αA-crystallin possesses high aggregation propensity and forms amyloid-like structures. αA(66-80) aggregates are known to interact with soluble crystallins and destabilize native structures that subsequently undergo aggregation. Crystallin aggregation in eye lens leads to reduction in lens opacity, the condition generally referred to as a cataract. Thus, αA(66-80) aggregation appears to be an important event during cataract development, and therefore, inhibition of αA(66-80) aggregation may be an attractive strategy to intervene in cataract development.αA(66-80) peptide derived from αA-crystallin possesses high aggregation potential and has a crucial role in cataract development. In order to inhibit the aggregation of αA(66-80) peptide, epigallocatechin-3-gallate (EGCG), a major active constituent of green tea, was employed. The inhibitory effect was assessed by Congo Red (CR) spectral shift assay, Thioflavin-T binding assay, transmission electron microscopy and fluorescence microscopy.The inhibitory potential of EGCG toward αA-crystallin was clearly observed as in the presence of EGCG, the αA(66-80) aggregation was considerably inhibited and the pre-formed fibrillary aggregates of αA(66-80) were found to be disassembled.In the present study, we are able to successfully demonstrate that EGCG efficiently blocks the aggregation of αA(66-80) peptide in a concentration-dependent manner. Furthermore, it is also evident that EGCG is able to disaggregate pre-formed αA(66-80) aggregates. The study suggests that EGCG can be a potential molecule that can prevent the initiation of cataract as well as be helpful in the disease reversal.

Seminal amyloids are well known for their role in enhancing HIV infection. Among all the amyloidogenic peptides identified in human semen, PAP248-286 was found to be the most active and was termed as semen-derived enhancer of viral infection (SEVI). Although amyloidogenic nature of the peptide is mainly linked with enhancement of the viral infection, the most active physiological conformation of the aggregated peptide remains inconclusive. Lipids are known to modulate aggregation pathway of a variety of proteins and peptides and constitute one of the most abundant biomolecules in human semen. PAP248-286 significantly differs from the other known amyloidogenic peptides, including Aβ and IAPP, in terms of critical concentration, surface charge, fibril morphology, and structural transition during aggregation. Hence, in the present study, we aimed to assess the effect of a lipid, 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), on PAP248-286 aggregation and the consequent conformational outcomes. Our initial observation suggested that the presence of the lipid considerably influenced the aggregation of PAP248-286 . Further, ZDOCK and MD simulation studies of peptide multimerization have suggested that the hydrophobic residues at C-terminus are crucial for PAP248-286 aggregation and are anticipated to be major DOPC-interacting partners. Therefore, we further assessed the aggregation behaviour of C-terminal (PAP273-286 ) fragment of PAP248-286 and observed that DOPC possesses the ability to interfere with the aggregation behaviour of both the peptides used in the current study. Mechanistically, we propose that the presence of DOPC causes considerable inhibition of the peptide aggregation by interfering with the peptide's disordered state to β-sheet transition.

The aim of the present study is to assess the effect of non-aqueous organic solvent on structural stability, molecular integrity and structure of α-amylase. The activity and thermal stability of the enzyme was measured before and after treatment with non polar solvent (i.e. hexane). The activity was found to be marginally affected and thermal stability was found to be significantly increased after treatment with hexane. The enzyme was found to be more resistant to thermal inactivation in hexane compared to in an aqueous buffer. The fluorescence measurement indicated a blue shift of 3 nm in the emission maximum ( λ max ) probably due to a minor change in the polarity of aromatic amino acid residues after treatment with a non-aqueous solvent. Assessment of thermal denaturation profile, 1-anilino-8-naphthalene-sulfonate (ANS) binding and acrylamide quenching of the enzyme suggested an increase in the molecular integrity and overall stability of the enzyme after treatment with hexane. However, these entire molecular events were not accompanied by any major change in the secondary structure. Our findings suggest that treatment of proteins or enzymes in non-aqueous solvents could be an attractive and cost-effective strategy to improve their structural stability without compromising their biological functions.

The effect of a cosolvent on the structure and stability of a protein depends upon the nature of preferential protein- water, protein-cosolvent or cosolvent-water interactions. The preferential interaction parameters of glycerol, sorbitol and sucrose with β-globulin (from Sesamum indicum L. seeds) were evaluated and the results showed the exclusion of cosolvents and preferential hydration of the protein. Data from fluorescence, circular dichroism (CD) and thermal stability measurements inferred that the preferential hydration had a considerable effect on the structure of protein under native conditions. Such cosolvent-protein interactions bring out a previously unnoticed, but outstanding phenomenon of cosolvent induced structural effects on the protein. This study reveals that these cosolvents interact with β-globulin in such a way that they induce a structural reshuffling to enhance the protein stability, mostly by intensifying intra-molecular hydrophobic interactions.

Since the discovery of Acquired Immuno Deficiency Syndrome (AIDS) in 1981 in United States, there have been tremendous efforts to reduce the rate of HIV transmission. Although, the epidemic is stabilized in most of the affected regions, its occurrence is reasonably evident in Eastern Europe and Central Asia due to high rate of new HIV infections. It is surprising to know that despite the high rate of infection, the virus is a weak pathogen. This paradox has been answered by a recent discovery stating that human semen contains a proteinaceous factor derived from prostatic acid phosphatase (PAP), which is commonly known as PAP248-286 peptide, plays an important role in enhancing the HIV infectivity. It forms well-defined amyloid structure, frequently referred as Semen-derived Enhancer of Viral Infection (SEVI) and enhances HIV infection up to 1,00,000 fold. Serendipitous discovery of this semen derived amyloid has provided an opportunity to design an alternative approach to dismantle the mechanism of HIV infection. It is a need of the hour to search and design novel molecules and compounds that can help in destabilizing SEVI under natural conditions. In this direction, a number of molecules have been identified that have shown promising results under laboratory conditions. However, there are several critical issues that remain untouched and their addressal is highly recommended in order to develop an effective regime to control the HIV transmission via sexual route. This review is an effort to consolidate major challenges in developing a therapeutic strategy against semen derived amyloids to combat HIV transmission.

The present study correlates the mechanism of α-amylase denaturation under acidic condition and its structural stabilization in presence of selected cosolvents (sorbitol, sucrose, trehalose, and glycerol). The objective of the present study was to minimize the enzyme inactivation at lower pH by stabilizing the enzyme structure. The above cosolvents were found to be an effective stabilizer of α-amylase against denaturation at extreme low pH. The optimum activity of α-amylase was found to be in the pH range of 4.5 to 7. Decreasing the pH of enzyme solution below this range results in a decrease in enzyme activity. The intrinsic and ANS fluorescence indicated the gradual unfolding of protein below pH 4 and resulted in exposure of hydrophobic cluster. The pH-induced unfolding also resulted in protein aggregation. The intrinsic and 8-anilinonaphathalene-1-sulphonic acid (ANS) fluorescence, acrylamide quenching, near and far UV-CD spectra of α-amylase at lower pH (pH 2.25) indicated the complete loss of tertiar...

N-terminal truncation and pyroglutamyl (pE) formation are naturally occurring chemical modifications of the Aβ peptide in Alzheimer's disease. We show herein that these two modifications significantly reduce the fibril length and the transition midpoint of thermal unfolding of the fibrils, but they do not substantially perturb the fibrillary peptide conformation. This observation implies that the N terminus of the unmodified peptide protects Aβ fibrils against mechanical stress and fragmentation and explains the high propensity of pE-modified peptides to form small and particularly toxic aggregates.

Alzheimer's disease (AD) is a fatal neurodegenerative disorder in humans and the main cause of dementia in aging societies. The disease is characterized by the aberrant formation of β-amyloid (Aβ) peptide oligomers and fibrils. These structures may damage the brain and give rise to cerebral amyloid angiopathy, neuronal dysfunction, and cellular toxicity. Although the connection between AD and Aβ fibrillation is extensively documented, much is still unknown about the formation of these Aβ aggregates and their structures at the molecular level. Here, we combined electron cryomicroscopy, 3D reconstruction, and integrative structural modeling methods to determine the molecular architecture of a fibril formed by Aβ(1-42), a particularly pathogenic variant of Aβ peptide. Our model reveals that the individual layers of the Aβ fibril are formed by peptide dimers with face-to-face packing. The two peptides forming the dimer possess identical tilde-shaped conformations and interact with each other by packing of their hydrophobic C-terminal β-strands. The peptide C termini are located close to the main fibril axis, where they produce a hydrophobic core and are surrounded by the structurally more flexible and charged segments of the peptide N termini. The observed molecular architecture is compatible with the general chemical properties of Aβ peptide and provides a structural basis for various biological observations that illuminate the molecular underpinnings of AD. Moreover, the structure provides direct evidence for a steric zipper within a fibril formed by full-length Aβ peptide.

In the present study an attempt was made to investigate the macromolecular crowding effect on functional attributes ofα-amylase. High concentrations of sugar based cosolvents, (e.g., trehalose, sucrose, sorbitol, and glycerol) were used to mimic the macromolecular crowding environment (of cellular milieu) underin vitroconditions. To assess the effect of macromolecular crowding, the activity and structural properties of the enzyme were evaluated in the presence of different concentrations of the above cosolvents. Based on the results it is suggested that the macromolecular crowding significantly improves the catalytic efficiency of the enzyme with marginal change in the structure. Out of four cosolvents examined, trehalose was found to be the most effective in consistently enhancing thermal stability of the enzyme. Moreover, the relative effectiveness of the above cosolvents was found to be dependent on their concentration used.

A differential relationship was observed between thermal stability and catalytic activity of α-amylase in the presence of different concentrations of CaCl(2). The enzyme displays optimum catalytic activity in the presence of 1.0-2.0 mM CaCl(2). Further addition of CaCl(2) leads to inhibition of the enzyme, however, at the same time the enzyme gains an additional resistance against thermal denaturation. It was evident that the enzyme is thermodynamically more stable (compared to the active enzyme) in the presence of inhibitory concentration of CaCl(2). For example, the thermal transition temperature (T(m)) of optimally active α-amylase was found to be 64±1°C, whereas, for the less active enzyme (in the presence 10 mM CaCl(2)) the value was determined to be 71±1°C. Similarly, the activation energy of thermal inactivation (Ea) was found to be 228±12 kJ/mol and 291±15 kJ/mol for the optimally active enzyme and the enzyme in the presence of 10 mM CaCl(2), respectively. Biophysical analysis of different states of the enzymes in response to variable calcium level indicates no significant change in the secondary structure in response to different concentration of CaCl(2), however the less active but thermodynamically stable enzyme (in the presence of higher concentration of CaCl(2)) was shown to have relatively more compact structure. The results suggest that the enzyme has separate catalytic and structure stabilizing domains and they significantly vary in their functional attributes in response to calcium level.

Amyloid oligomers are nonfibrillar polypeptide aggregates linked to diseases, such as Alzheimer's and Parkinson's. Here we show that these aggregates possess a compact, quasi-crystalline architecture that presents significant nanoscale regularity. The amyloid oligomers are dynamic assemblies and are able to release their individual subunits. The small oligomeric size and spheroid shape confer diffusible characteristics, electrophoretic mobility, and the ability to enter hydrated gel matrices or cells. We finally showed that the amyloid oligomers can be labeled with both fluorescence agents and iron oxide nanoparticles and can target macrophage cells. Oligomer amyloids may provide a new biological nanomaterial for improved targeting, drug release, and medical imaging.

Accumulation of ordered protein aggregates (or amyloids) represents a hallmark of many diseases (e.g., Alzheimer's disease, type II diabetes, Parkinson's diseases etc.), results from intermolecular association of partially unfolded proteins/ peptides. Such associations usually take place in highly crowded conditions. The aggregates, which are formed under in vitro and in vivo conditions exhibit substantial variations in their structure and function. Such heterogeneities in amyloids might arise due to macromolecular crowding that is usually omitted under in vitro conditions. The current study is an attempt to assess the effects of macromolecular crowding on amyloid formation using a model amyloidogenic peptide. The sequence of the peptide was derived from C-terminal region (RATQIPSYKKLIMY) of PAP(248-286), which naturally occurs in human semen as amyloid aggregates and is known for boosting HIV infectivity. This model peptide forms sedimentable and fibrillar aggregates in aqueous buffer and shows the characteristic features of amyloids. In the presence of macromolecular crowders the morphological features of the amyloids are significantly altered and resulted in the formation of shorter amyloid aggregates. The current study assumes the hypothesis that macromolecular crowding in the biological system favours formation of heterogeneous classes of aggregates and each of them might differ in their biophysical and biological properties.

Naturally occurring fragments of the abundant semen proteins prostatic acid phosphatase (PAP) and semenogelins form amyloid fibrils in vitro. These fibrils boost HIV infection and may play a key role in the spread of the AIDS pandemic. However, the presence of amyloid fibrils in semen remained to be demonstrated. Here, we use state of the art confocal and electron microscopy techniques for direct imaging of amyloid fibrils in human ejaculates. We detect amyloid aggregates in all semen samples and find that they partially consist of PAP fragments, interact with HIV particles and increase viral infectivity. Our results establish semen as a body fluid that naturally contains amyloid fibrils that are exploited by HIV to promote its sexual transmission.

Aggregation of b-amyloid (Ab) peptide into oligomers and protofibrils is a hallmark of Alzheimer s disease (AD). Increasing evidence shows that the primary insult in AD is caused by oligomeric species that impair the ordered function of synaptic networks. Consistent with this view, oligomers were shown to affect synaptic plasticity, and they impair the long-term potentiation (LTP) in living brain tissues, a widely used model system of brain memory functions. Using solidstate NMR spectroscopy, we here determined the residuespecific molecular conformation of a highly synaptotoxic bamyloid oligomer structure. Our measurements reveal a stable N-terminal b strand that controls the partitioning between oligomer and protofibril formation, whereas targeting the peptide N-terminus ameliorates Ab-dependent neuronal dysfunctions. The presently investigated, chemically well-defined Ab oligomers faithfully reproduce the hallmark characteristics of AD-related oligomers. Living hippocampal brain slices were exposed to different Ab conformers (Figure 1A), and a series of tetanic electrical stimuli were applied to evoke a longlasting increase of the synaptic transmission, termed LTP. Oligomers, but not freshly dissolved, that is, primarily monomeric, Ab peptide or fibrils, reduce the LTP response and therefore disturb the brain memory functions within these tissue samples (Figure 1B). A similar oligomer-specificity is seen with cultured primary neurons, which present a significant oligomer-dependent decrease ( 40%) of their

The present paper is an attempt to study the mechanism of ethanol induced aggregation of chicken egg albumin and to stabilize the protein against ethanol induced aggregation. The protein aggregation was determined by monitoring the light scattering of protein aggregates spectrophotometrically. The protein undergoes certain structural changes in water- ethanol solution and the degree of aggregation was found to be linearly depending upon the concentration of alcohol used. The intrinsic fluorescence study showed a large blue shift in the λmax (16 nm) in the presence of 50% ethanol. The ANS fluorescence intensity was found to be gradually increasing with increasing concentration of ethanol. This indicates an increase in the hydrophobic cluster on the protein surface and as a result the hydrophobic interaction is appeared to be the major driving force for the aggregate formation. Addition of sucrose significantly reduced the ethanol-induced aggregation of chicken egg albumin. In presence of 50% sucrose the ethanol induced aggregation was reduced to 5%. The study reveals that addition of sucrose brings out changes in the solvent distribution and prevents the structural changes in protein which lead the aggregation.

The activity and thermal stability of alpha-amylase were studied in the presence of different concentrations of trehalose, sorbitol, sucrose and glycerol. The optimum temperature of the enzyme was found to be 50 +/- 2 degrees C. Further increase in temperature resulted in irreversible thermal inactivation of the enzyme. In the presence of cosolvents, the rate of thermal inactivation was found to be significantly reduced. The apparent thermal denaturation temperature (Tm) app and activation energy (Ea) of alpha-amylase were found to be significantly increased in the presence of cosolvents in a concentration-dependent manner. In the presence of 40% trehalose, sorbitol, sucrose and glycerol, increments in the (Tm)app were 20 degrees C, 14 degrees C, 13 degrees C and 9 degrees C, respectively. The Ea of thermal denaturation of alpha-amylase in the presence of 20% (w/v) trehalose, sorbitol, sucrose and glycerol was found to be 126, 95, 90 and 43 kcal/mol compared with a control value of 40 kcal/mol. Intrinsic and 8-anilinonaphathalene-1-sulphonic acid (ANS) fluorescence studies indicated that thermal denaturation of the enzyme was accompanied by exposure of the hydrophobic cluster on the protein surface. Preferential interaction parameters indicated extensive hydration of the enzyme in the presence of cosolvents.