The problem tested in the experiment is whether the protein-osmolyte-water solution stabilizes or destabilizes the unfolded state of protein. This problem was used to test the mechanism by which the osmolytes, the small organic compounds, interact with the protein to affect stability. The proposed hypothesis stated that the transfer free energy of protein backbone form water to a water/osmolyte solution, Δgtr, is negatively correlated with an osmolyte’s fractional polar surface area. Δgtr is the unit used to measure the degree to which an osmolyte stabilizes the protein given that if Δgtr>0 (stabilized) and Δgtr<0 (destabilized). The independent variable was the fractional polar surface area (SA) and the dependent variable was the energy change or stability (Δgtr). The model had other two adjustable parameters or independent variables: the polar and non-polar SAs and the interaction between solvent and the protein backbone. The Δgtr was calculated using all the adjustable parameters. The first experiment involved the quantitative solvation model in which the solvent interaction energy is the function of interactant polarity. The other experiment involved the number of energetically equivalent ways of realizing a given interaction is a function of interactant surface area. To perform the experiment researchers used 1 M osmolyte solution. Using x-ray structure of eight stabilizing osmolytes calculations were performed. Even though the comparisons between the osmolyte structure and the Δgtr values indicate no evident correlation, there was a clear correlation between Δgtr and fractional surface polar area (R= 0.88). The second experiment used the quantitative model for solvent (water and osmolyte) to test the interactions with backbone polar groups. It indicated that the back-bone/osmolyte interactions became increasingly favorable as osmolyte became increasingly polar. The result stated for 1 M osmolyte concentration, the calculated and the measured Δgtr values are in good agreement. Especially as the fractional surface polar area increases, the osmolyte interaction with the protein backbone becomes increasingly favorable, that is their Δgtr values decreases. The free energy change for folding/unfolding will be linearly dependent on osmolyte concentration. The statistical mechanics model was used to calculate the average energy of the protein backbone in osmolyte solutions. Approximately 90% calculated Δgtr have the correlation exceeding 0.80. The success of the second experiment was consistent with their hypothesis.
The single amide nitrogen and two carbonyl oxygen backbone interactions were used for solvent interactions. Each of these sites had ether positive, neutral or negative charge presented by the solvent (water or osmolyte).
Abstract/Summary: “Proteins account for more than 50% of the dry weight of most cells, and they are instrumental in almost everything organisms do” (Campbell, 1999). The significance of proteins to the continuation of our biological systems is undeniable, and a study of how to quantify proteins seems an appropriate introduction to our studies of biology. In order to study proteins we must first know how to separate then quantify the amount using basic principles of experimental design such as a standard curve. In this experiment we wish to quantify the amount of previously extracted protein by measuring the absorbance of the unknown amount and determining its concentration by overlaying it against a standard curve of the absorbance of known concentrations of the protein. We used the dye agent Bradford Protein Assay to get an absorbance of 0.078, 0.143, 0.393, 0.473, and 0.527 at the protein’s respective concentrations of 0.28, 0.56, 0.84, 1.12, and 1.40 mg/mL. When a best-fit line was applied to the standard curve, and the absorbance of our unknown concentration (0.317 A) plotted, we estimated a concentration of around 0.84 mg/mL of protein. Our calculations indicated a quantity of 168 mg of protein, which was an approximately 8.96% yield of the projected 1875 mg that was expected. Errors that may have led to this small yield percentage may have stemmed from our previous lab and our initial attempts to extract the desired amount of protein.
This experiment focuses on the SN2 nucleophile substitution reaction of converting 1-butanol (an alcohol) to 1-bromobutane (an alkyl halide). There are two types of substitution mechanisms that could be used, SN1 and SN2. SN1 mechanisms take place in two steps. The first rate-determining step is the ionization of the molecule. This mechanism is called unimolecular because its rate is only dependent on the concentration of the leaving group. The second step is the fast, exothermic nucleophile addition. In an SN2 reaction the leaving group leaves as the nucleophile attacks all in one step. Because this happens at one time, the nucleophile must attack from the opposite side from which the leaving group is leaving. For this reason, SN2 reactions
Sequence and structural proteomics involve the large scale analysis of protein structure. Comparison among the sequence and structure of the protein enable the identification on the function of newly discovered genes (Proteoconsult, n.d.). It consists of two parallel goals which one of the goals is to determine three-dimensional structures of proteins. Determine the structure of the protein help to modeled many other structures by using computational techniques (Christendat et al., 2000). This approach is useful in phylogenetic distribution of folds and structural features of proteins (Christendat et al., 2000). Nuclear magnetic resonance (NMR) spectroscopy is one of the techniques that provide experimental data for those initiatives. It is best applied to proteins which are smaller than 250 amino acids (Yee et al., 2001). Although it is limited by size constraints and also lengthy data collection and analysis time, it is still recommended as it can deliver strong results. There are two types of NMR which are one-dimensional NMR and two-dimensional NMR. One-dimensional NMR provides enough information for assessing the folding properties of proteins (Rehm, Huber & Holak, 2002). It also helps to identify a mixture of folded and unfolded protein by observing both signal dispersion and prominent peak. Observation in one-dimensional spectrum also obtains information on molecular weight and aggregation of molecule under investigation. In spite of this, two-dimensional NMR are used for screening that reveal structural include binding, properties of proteins. It also provides important information for optimizing conditions for protein constructs that are amenable to structural studies (Rehm et al., 2002). NMR is a powerful tool which it w...
The three-dimensional contour limits the number of substrates that can possibly react to only those substrates that can specifically fit the enzyme surface. Enzymes have an active site, which is the specific indent caused by the amino acid on the surface that fold inwards. The active site only allows a substrate of the exact unique shape to fit; this is where the substance combines to form an enzyme- substrate complex. Forming an enzyme-substrate complex makes it possible for substrate molecules to combine to form a product. In this experiment, the product is maltose.
According to osmosis theory as the concentration of the sucrose solution increases the particles water potential increases and becomes higher than the particles that are in the
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The Effect of pH on Enzyme Activity. pH is a measure of the concentration of hydrogen ions in a solution. The higher the hydrogen ion concentration, the lower the pH. Most enzymes function efficiently over a narrow pH range. A change in pH above or below this range reduces the rate of enzyme reaction. considerably.
The covalent structure of a protein is composed of hundreds of individual bonds. Because free rotation is possible around a good portion of these bonds, there are a very high number of possible conformations the protein can assume. However, each protein is responsible for a particular chemical or structural function, signifying that each one has a distinctive three-dimensional configuration. By the early 1900’s, numerous proteins had been crystallized. Because the ordered collection of molecules in a crystal can only form if all of the molecular units are the same, the discovery that proteins could be crystallized proved that even large proteins have distinct chemical structures. This deduction completely transformed the understanding of proteins and their respective functions. It is important to investigate how a series of amino acids in a polypeptide chain is translated into a three-dimensional protein structure. There are five general topics related to this process: the structure of a protein is determined by its amino acid sequence, the role of a protein is dependent on its unique structure, an isolated protein typically exists in a small number of stable forms, non-covalent interactions are the most important stabilizing forces in a protein structure, and there are structural patterns that aid in explaining and understanding protein architecture.
The majority of the chemical reactions associated with diphenyl ether relate to the structures of the two phenyl rings of the organic compound due to the lack of reactivity typically associated with ethers. Ethers usually make excellent solvents due to the characteristic lack of reactivity. However, diphenyl ether participates in interesting reactions in spite of the stereotype. First and foremost, the formation of diphenyl ether primarily results from a deviation of Williamson Ether synthesis. These general reactions “involve an alkoxide that reacts with a primary haloalkane or a ...
The Functions of Proteins Introduction Protein accounts for about three-fourths of the dry matter in humans. tissues other than fat and bone. It is a major structural component of hair, skin, nails, connective tissues, and body organs. It is required for practically every essential function in the body. Proteins are made from the following elements: carbon, hydrogen, oxygen, nitrogen. and often sulphur and phosphorus.
Proteins are large molecules that play an integral role in the body’s function. Proteins perform functions in the body such as enzyme catalysis, DNA replication, cell signaling, and transportation of molecules from one location to another. Proteins are made up of smaller units called amino acids, which are made from the 20 amino acids. What makes proteins differ from one another is the specific sequence of amino acids and their three-dimensional structure. There are four distinct structures a protein can have which are primary, secondary, tertiary, and quaternary. As proteins begin to form during the primary stage they start out in a linear chain of amino acids. In the secondary structure the linear chain of amino acids begins to twist. In the tertiary structure the amino acid chains continue to fold and twist and form bonds from disulfide bridges, which are made of two sulfur atoms. In the final and quaternary structure the chains fold together into a tighter knit structure forming proteins such as hemoglobin.
An enzyme is a specialized protein that acts as a catalyst and facilitates complex metabolic processes. An enzyme, like any protein, is a polymer made up of a long chain of amino acids. The sequence of amino acids is determined by the DNA template in which it was made, and the amino acids are attached together by peptide bonds. Cross linking takes place between the R groups of the amino acids and forms a unique three dimensional molecules. The structure and spatial configuration of an enzyme, especially its binding site, is key to its optimal function and activity. This 3-dimensional structure can easily be altered by environment factors, such as salinity and pH. Each enzyme has a binding site in which chemical bonds are achieved with their
Proteins are considered to be the most versatile macromolecules in a living system. This is because they serve crucial functions in all biological processes. Proteins are linear polymers, and they are made up of monomer units that are called amino acids. The sequence of the amino acids linked together is referred to as the primary structure. A protein will spontaneously fold up into a 3D shape caused by the hydrogen bonding of amino acids near each other. This 3D structure is determined by the sequence of the amino acids. The 3D structure is referred to as the secondary structure. There is also a tertiary structure, which is formed by the long-range interactions of the amino acids. Protein function is directly dependent on this 3D structure.
There are four main levels of a protein, which make up its native conformation. The first level, primary structure, is just the basic order of all the amino acids. The amino acids are held together by strong peptide bonds. The next level of protein organization is the secondary structure. This is where the primary structure is repeated folded so that it takes up less space. There are two types of folding, the first of which is beta-pleated sheets, where the primary structure would resemble continuous spikes forming a horizontal strip. The seco...
J. Clayden, N. Greeves, S. Warren, P. Wothers. Organic Chemistry. 8th ed. 2007, Oxford University Press, p. 1186-1191.