Sharif Digital Repository

At the molecular level, proteins control almost all the biochemical reactions in the cells. In order for proteins to function, they must be able to fold into their unique 3D structure. Deformation of protein structure due to some environmental phenomena such as pH, high temperature, and stress may cause unfolding of proteins and finally result in formation of protein fibrils called "amyloid". Amyloid has been found in a number of human diseases such as Alzheimer, type II diabetes, and Parkinson. Recently pyridazine has been under spotlight due to its unique chemical properties such as high dipole moment and higher solubility in biological solvents. In this study, we aim at using a... 

Bioorthogonal chemistry defines any reaction, inside or outside the living systems, without interfering with native chemical processes and 3dimensional structure of proteins. In these types of reactions water is the sole solvent; a neutral pH is required; the temperature must be up to 40 oC. The kinetics of reactions must be on the hour scale and the nontoxic reagents with low concentration must be used. One of the simplest methods of visualizing protein molecule is covalent attachment of fluorescein to protein. In this work several fluorescent compounds were synthesized. These novel compounds include acylchloride fluorescein, N-hydroxysuccinimide fluorescein, thiophenol fluorescein, and... 

The most important factor in protein's function and its 3D structure is folding. Misfolding or native folding perturbation in some peptides and proteins force them get into amyloid fibril aggregation. It has been observed that these aggregations are the cause of some debilitative diseases especially ones that are related to aging like Alzheimer disease, Hantington's disease, typeII diabets etc. Recently it has been shown that specific regions in human catalase protein interact with some amyloid fibrils, such as Aß, IAPP, PrP etc. Inteins are self-splicing protein enzymes, which excise themselves from the mature protein and join the two flanking adjacent sequences by a new peptide bond in a... 

Transglutaminase 2 enzyme is expressed in various parts of the human body and is present in almost all tissue cells. The function of this enzyme is multiple and it catalyzes different reactions depending on where it is located in the cell. The most important activities of this enzyme are: isopeptide bond formation between the side chains of two proteins (transamidation reaction), diamidation reaction (hydrolysis of amide bond), esterification, kinase activity. The hen egg white lysozyme protein is another natural antimicrobial protein that has been extensively studied. The muramidase activity of egg white lysozyme against highly gram-positive bacteria is well known, and this property has... 

The reductase enzymes perform an important function in the cell. These enzymes are usually coupled with other enzymes and participate in reactions such as halogenation, reduction of alkene, and carboxylic acids. These reactions are carried out by reproducing cofactors such as FADH2 and NAD+; however, there is no evidence that this enzyme alone can perform an organic reaction. In this work, we aim to examine whether recombinant flavin reductase can carry out organic reactions. In order to amplify the flavin reductase enzyme gene from the E. coli genome by PCR (Polymerase Chain Reaction) method, two synthetic primers were designed; The forward primer of the gene contained a cleavage site for... 

Quorum Sensing is a cell-to-cell communication pr°Cess depending on cell concentration. Communication among bacterial cells °Ccurs through small molecular signals called autoinducers. The consequences of quorum sensing include the activation of pathogenic properties and activation of the gene for biofilm formation. Biofilms are aggregations of bacteria that adhere to surfaces through extracellular polymeric substances. Bacteria in the biofilm phase are much more resistant to antimicrobial agents than their free-swimming counterparts (i.e. the bacteria in planktonic form). In this study, first a series of coumarin derivatives were synthesized; but due to low solubility of the synthesized... 

Asparagainase is a therapeutic enzyme which has been a subject of research for decades. The enzyme catalyzes the hydrolysis of the amide group in asparagine and similar amides. Altering the substrate specificity and stabilization of this enzyme can increase its therapeutic properties. Moreover, asparaginases may be evolved to catalyze the hydrolysis of other similar compounds. These can be achieved through directed evolution and computational methods.In this study, the gene encoding L-asparaginase II enzyme from E. coli was amplified by polymerase chain reaction (PCR) and was cloned into an expression vector. The recombinant protein was expressed by an appropriate host secreting the... 

Red phosphorus, due to its unique chemical properties, and glycidol, with its reactive functional groups, provide an effective combination for synthesizing new copolymers with potential applications in biological fields, particularly in enzyme stabilization. In this study, a novel copolymer was synthesized using red phosphorus and glycidol through an anionic ring-opening mechanism. The synthesis process was confirmed using various analytical techniques, including FTIR, NMR, and thermal analyses, which demonstrated the successful incorporation of functional groups and an average phosphorus content of 40% in the copolymer. In the next step, modification of this copolymer by linking it to the... 

Carbon Quantum Dots (CQDs) are a new class of zero-dimensional carbon-based nanoparticles with a size range from 1 to 10 nanometers. In this study, it was tried to form new carbon-nitrogen bond in organic synthesis using non-metal CQD-catalysis, thereby taking a significant step towards enhancing the catalytic role of carbon nanoparticles. Carbon-nitrogen bond is of paramount importance in biology and organic chemistry. In this regard transamidation reaction, reductive amination, and thioureas syntheses as three important reactions in organic chemistry were explored using non-metal CQD-catalysis. Transamidation is mainly used to produce new amides by forming carbon-nitrogen bond. This... 

Enzymes are proteins that carry out chemical reactions in the form of chemo, stereo, and regioselective under mild conditions. One of the characteristics of enzymes is that they perform a specific chemical reaction on a selective substrate and under specific environmental conditions. However, in recent years, a number of enzymes have been identified to display promiscous behavior. This promiscousity includes catalytic activity, variety of substrates, and environmental conditions. Promiscous activities of enzymes can be a good starting point for creating new and more favorable engineered pathways for the production of chemical and medicinal drugs. One of the aims of this project is to exploit... 

Proteins carry out most important roles in the organisms and they have a number of functions including structural, catalytically, regulatory and transportal, However for protein to have functional role they must be able to possess tridimensional structure. Several proteins due to various genetic and physico-chemical conditions have shown to alter their tridimensional structure leading to insoluble fibrilar structure. One of the properties of these insoluble fibrils is conformational change, converting to β-sheet rich structures; the entire process of converting soluble to proteins insoluble fibrils known as amyloid formation. The proteins amyloidosis have been seen as causative role in a... 

Nowdays, it has been shown that protein misfolding is involved in more than forty diseases including Alzheimer’s and Parkinson’s. Alzheimer’s disease is one of the most common neurodegenerative diseases that disrupts brain function. The main cause of this illness is due to formation of extracellular amyloid plaque from a 42-amino acid-peptide called amyloid beta. Amyloidosis is caused by missfolding of peptides and proteins mainly because of physical and chemical changes in biomolecule or their environment.
Therefore, in this dissertation, using synthetic and recombinant abeta peptide, kinetic of amyloidosis was examined. Ultrasmall peptides (three-amino acid), containing branched chain... 

Transglutaminases of family enzymes are implicated in different functions in cell including cellular division and cell adhesion. Several members of this family catalyze formation of isopeptide bond that involved in wound restoration and angiogenesis. Transglutaminase 2 that have been studied in this project, are found in almost every human cell tissue. This enzyme has multiple functions depending on its cell location. In cytoplasm TG2 act as GTPase and is involve in post translational modifications. This enzyme by using calcium form a isopeptide bond between two proteins, isopeptide bond is function in brain cortex and cells creates amyloid structures that including brain diseases, Parkinson... 

Due to the importance of using pure proteins in the industry and basic research in order to study them in the structural contexts and interactions of proteins in recent years, purification methods have been extensively developed. The purification steps usually depend on the size of the protein, its physical and chemical properties, the binding affinity and the biological activity of the protein in question, In the purification process, the protein portion is separated from the non-protein portion. The biggest challenge is when we want to separate different proteins. Different chromatographic methods are used, depending on the properties of the impure proteins. One of these chromatographic... 

After proteins, one of the important biological macromolecules, are synthesized by ribosomes,they automatically reach their own special 3-D structure. This native structur or better known as folded state is what determines the function of a protein. In some situations, the proteins become misfolded leading to its lack of biological function through aggregation or fibril formation known as "amyloid". The amyloid fibrils have been shown to be a causative factor in nerve decay such as Alzheimer's. Therefore, inhibition of amyloid formation using different approaches such as synthetic organic and natural compounds has, recently, been under spotlight. A pyridazine basedcompound (RS-0406) has... 

Hyperbranched copolymers are highly attractive due to their unique properties, such as tunable structure and surface functionality, making them versatile for applications in biotechnologies, pharmaceuticals, and materials engineering. Glycidol, as a reactive monomer with excellent biocompatibility, combined with sulfur, which enhances mechanical and chemical properties, represents a promising combination for hyperbranched copolymer synthesis. In this study, hyperbranched glycidol-sulfur copolymers were synthesized using cationic and anionic ring-opening polymerization methods. FT-IR analysis confirmed the successful incorporation of S-S bonds and sulfur-containing functional groups into the... 

Due to its structural simplicity and functional importance, Hen egg white lysozyme (HEWL) is a widely studied model protein. However, its expression in E. coli often results in aggregation and the formation of inclusion bodies, limiting its utility for biotechnological applications. This study focuses on improving the solubility and stability of HEWL through a combination of experimental and computational approaches. In the first step, using error-prone PCR and the MEGAWHOP technique, libraries of potential HEWL variants were generated and screened via β-complementation for enhanced properties. To complement these efforts, computational tools such as ProteinMPNN and AlphaFold were employed... 

Enzymes are biological catalysts that carry out chemical reactions in a stereoselective manner. Fumarate hydratase (abbreviated as fumarase) is a metabolic enzyme found in both mitochondrial and cytosolic isoforms. Its mitochondrial form functions in the Krebs cycle (TCA) and specifically converts fumarate into L-malate. In recent studies, the role of the cytosolic form of this enzyme in the repair of DNA double-strand breaks has been elucidated. Fumarase and its precursors are indirectly related to many metabolic pathways, including phenylalanine and tyrosine metabolism, urea cycle, etc. As a result, it is very important to know the factors affecting this enzyme and its activity. In this... 

L-asparaginase II is an amidohydrolase which hydrolyzes L-asparagine to furnish L-aspartic acid and ammonia. The enzyme has played an important role for 40 years in treating cancer in persons with juvenile leukemias or lymphomas. Furthermore, the enzyme has become important in food industries and biosensors technology.In this study, the expression of recombinant L-asparaginase II in E. Coli was optimized and the purification of this enzyme from growth culture was carried out with nickel-charged affinity resin. Afterwards, the activity, and kinetic parameters of purified recombinant L-asparaginase II including Km, kcat, Vmax, and enzyme efficiency were measured.In the next step, random... 

The main purpose of this Ph.D. Thesis is to develop DNA-based biosensors using Label-free approaches applying simple, inexpensive and fast electrochemical techniques to measure the biological markers of cancer.In the first part, a simple protocol for detection of specific-sequence DNA is introduced. In this method carbon nanotube is used as a hybridization indicator. This label-free system provides the advantage of eliminating additional labeling procedure. As the signal enhances in the presence of MWCNT and decreases in the presence of target, the fabricated sensor is known as a signal-off device. The oxidation signal of Fe(CN)63-/4- is followed as an analytical signal to detect target...