Ashim Paul

Reversal of aggregation using β-breaker dipeptide containing peptides: Application to Aβ(1-40) self-assembly and its inhibition.

Reversion of protein or peptide aggregation is a formidable task, important in various domains of research at the interface of chemistry, medicine, and nanoscience. A novel class of dipeptides, termed as β-breaker dipeptides (BBDPs), is identified, which can be incorporated into the self-recognizing sequences to generate a novel class of conformational switch which forms β-sheet at an initial stage and then converts in a controlled manner to random coil at specific conditions. Incorporation of BBDPs in a well designed amyloidogenic peptides generates a special class of β-sheet breaker peptides those undergo a chemical change at physiological condition generating a breaker element in situ. These β-breaker peptides are shown to first incorporate into the amyloid and then disrupt it. Such conformational switches may be used to study agrregation/disaggregation process and may find many biomedical applications relevant to aggregation related disorders. Such strategy for reversion of peptide aggregation using chemical tricks may find application in material chemistry as well.

Waste reduction in amide synthesis by a continuous method based on recycling of the reaction mixture

In this article, a new one pot synthesis of amides is demonstrated. Potential of the recovery and reuse strategy for waste reduction is examined. Finally, a new strategy is proposed where the whole reaction mixture containing the catalyst, the solvent (auxiliary) and the unreacted starting material is recovered by precipitating the product using ethanol and reused for the next cycle in a continuous operation protocol.

Disaggregation of Amylin Aggregate by Novel Conformationally Restricted Aminobenzoic Acid containing α/β and α/γ Hybrid Peptidomimetics

Diabetes has emerged as a threat to the current world. More than ninety five per cent of all the diabetic population has type 2 diabetes mellitus (T2DM). Aggregates of Amylin hormone, which is co-secreted with insulin from the pancreatic β-cells, inhibit the activities of insulin and glucagon and cause T2DM. Importance of the conformationally restricted peptides for drug design against T2DM has been invigorated by recent FDA approval of Symlin, which is a large conformationally restricted peptide. However, Symlin still has some issues including solubility, oral bioavailability and cost of preparation. Herein, we introduced a novel strategy for conformationally restricted peptide design adopting a minimalistic approach for cost reduction. We have demonstrated efficient inhibition of amyloid formation of Amylin and its disruption by a novel class of conformationally restricted β-sheet breaker hybrid peptidomimetics (BSBHps). We have inserted β, γ and δ -aminobenzoic acid separately into an amyloidogenic peptide sequence, synthesized α/β, α/γ and α/δ hybrid peptidomimetics, respectively. Interestingly, we observed the aggregation inhibitory efficacy of α/β and α/γ BSBHps, but not of α/δ analogues. They also disrupt existing amyloids into non-toxic forms. Results may be useful for newer drug design against T2DM as well as other amyloidoses and understanding amyloidogenesis.

Amyloid β derived switch-peptides as a tool for investigation of early events of aggregation: a combined experimental and theoretical approach

Alzheimers disease, a severe neurodegenerative disorder, is believed to be caused by the interneuronal aggregation of the amyloid β peptide. It has no cure yet. Despite rigorous research, the mechanism of aggregation is not yet fully delineated. Especially probing the early events of the aggregation is difficult as we have no control on the kinetics of the process of aggregation. We have used amyloid β derived switch-peptides that behave as the functional mimic of the amyloid β peptide, and demonstrated that side chain aromatic interactions precede the β-sheet formation resulting in fibrillization. Detailed investigation into the early events of aggregation has become possible as the kinetics of aggregation of the switch-peptides can be controlled. We have used ultraviolet (UV) spectroscopy, circular dichroism (CD), Raman spectroscopy and Molecular Dynamics (MD) simulation as a combined experimental and theoretical approach.

Protective effects of β-sheet breaker α/β-hybrid peptide against amyloid β-induced neuronal apoptosis in vitro.

Alzheimers disease is most common neurodegenerative disorder and is characterized by increased production of soluble amyloid‐β oligomers, the main toxic species predominantly formed from aggregation of monomeric amyloid‐β (Aβ). Increased production of Aβ invokes a cascade of oxidative damages to neurons and eventually leads to neuronal death. This study was aimed to investigate the neuroprotective effects of a β‐sheet breaker α/β‐hybrid peptide (BSBHp) and the underlying mechanisms against Aβ40‐induced neurotoxicity in human neuroblastoma SH‐SY5Y cells. Cells were pretreated with the peptide Aβ40 to induce neurotoxicity. Assays for cell viability, cell membrane damage, cellular apoptosis, generation of reactive oxygen species (ROS), intracellular free Ca2+, and key apoptotic protein levels were performed in vitro. Our results showed that pretreatment with BSBHp significantly attenuates Aβ40‐induced toxicity by retaining cell viability, suppressing generation of ROS, Ca2+ levels, and effectively protects neuronal apoptosis by suppressing pro‐apoptotic protein Bax and up‐regulating antiapoptotic protein Bcl‐2. These results suggest that α/β‐hybrid peptide has neuroprotective effects against Aβ40‐induced oxidative stress, which might be a potential therapeutic agent for treating or preventing neurodegenerative diseases.

The role of caffeine as an inhibitor in the aggregation of amyloid forming peptides: a unified molecular dynamics simulation and experimental study

Alzheimer’s disease is a devastating neurodegenerative disease triggered by the aggregation of amyloid-β peptide (Aβ) into amyloid fibrils. From the results of several studies it is believed that caffeine can prevent the development of Alzheimer’s disease. However, the molecular mechanism of the therapeutic potential of caffeine is largely unknown. In our study, we have investigated the effect of caffeine on the aggregation of amyloid-β derived switch-peptide by varying the stoichiometric ratio of caffeine to peptide. Our molecular dynamics study of peptides in pure water show the formation of a β-sheet conformation, which is prevented to a large extent in the presence of a 10 : 1 or greater ratio of caffeine to peptide. The experimental results demonstrate that caffeine can inhibit the formation of β-sheets by interacting with the peptide aromatic moiety. A detailed molecular dynamics analysis of the inhibition of peptide aggregation by caffeine further revealed that caffeine molecules form hydrogen bonds with peptides thereby weakening the interstrand hydrogen bonds between the peptides. The self-aggregated caffeine clusters form a hydrophobic environment around the hydrophobic residues of the peptides, and physically block them from interacting with each other.

Phenolic Ester Mediated Oligopeptide Synthesis Promoted by HOBt

Although substituted phenolic ester mediated peptide synthesis is an efficient and well established method, the same via totally unsubstituted phenyl ester is not preferred due to the extremely slow rate of aminolysis. We have investigated the scope of the unsubstituted phenyl ester as an intermediate in peptide bond formation and found that it may be useful for the design of chemoselective peptide ligation when HOBt is used as an acyl transfer catalyst. The scope of HOBt catalyzed, oxo ester mediated ligation is explored for the synthesis of oligopeptides containing a cysteine, serine and threonine at the N-terminus of the ligating peptide.

A Peptide Based Pro-drug Disrupts Alzheimer’s Amyloid into Non-toxic Species and Reduces Aβ Induced Toxicity In Vitro

Aggregation of Amyloid β (Aβ) in the interneuronal spaces is a major etiopathological factor for onset and progression of Alzheimer’s disease (AD). Since the mechanism of aggregation is not fully understood, control and modulation of the aggregation process is a challenging task. Although, several strategies were developed for the past few decades, yet there is no proper therapeutics available. Herein, we report a peptide based pro-drug, termed as a conformational Pro-Drug peptide (PDp), which disrupts existing Aβ fibrils, but does not produce toxic soluble oligomers, through a series of spontaneous chemical reactions resulting in in situ generation of β-sheet destabilizing factors. Furthermore, PDp reduces Aβ mediated toxicity examined on an in vitro model consisting of the human neuroblastoma SH-SY5Y cells. PDp also disrupts fibrils originated from AD affected human cerebrospinal fluid. These findings will help to understand the process of amyloidogenesis better and also indicate a novel approach for therapeutically important peptide design.

A Peptide Based Pro-Drug Ameliorates Amyloid-β Induced Neuronal Apoptosis in in vitro SH-SY5Y cells.

BACKGROUND Alzheimers disease (AD), a common protein misfolding progressive neurodegenerative disorder, is one of the most common forms of dementia. Amyloid precursor protein (APP) derived amyloid-β (Aβ) protein accumulate into interneuronal spaces and plays a crucial role in the disease progression and its pathology. The aggregated Aβ exerts its neurotoxic effects by inducing apoptosis and oxidative damage in neuronal cells. OBJECTIVES We have investigated the effects of a synthesized Pro-Drug peptide (PDp) on Aβ1-40 induced cytotoxicity in human neuroblastoma SH-SY5Y cells, represents one of the most effective strategies in combating human AD. METHODS Cells were treated with Aβ1-40 to induce cytotoxicity in the experimental model of AD to screen the inhibitory effect of PDp. Assays for cell viability, reactive oxygen species (ROS) generation, levels of intracellular free Ca2+ and expression of key apoptotic proteins were assessed by Western Blotting. RESULTS Our results showed that Aβ1-40 induces for 24h caused reduce cell viability, imbalance in Ca2+ homeostasis and increase in neuronal apoptosis in vitro. Treatment with PDp could effectively ameliorated Aβ1-40 induced neurotoxicity and attenuates ROS generation that mediates apoptotic signaling through Bcl-2, Bax, Caspase-3 activity and cytochrome c in the cells. CONCLUSION These findings suggested that PDp has potential role as a neuroprotective and therapeutic agent for combating human AD.

Microwave assisted chemoselective organocatalytic peptide alcohol synthesis from C-terminal amide

The synthesis of peptide alcohols from peptides bearing C-terminal amide using β-aminoalcohols is achieved by a simple, efficient, racemization-free (<1.5%), and chemoselective transformation in the presence of cost-effective PTSA (para-toluenesulfonic acid) under microwave irradiation. Peptide alcohols can be synthesized by a standard solid phase peptide synthesis protocol obviating the need for specially designed resin.