An oxyanion hole is a pocket in the active site of an enzyme that stabilizes transition state negative charge on a deprotonated oxygen or alkoxide. The pocket typically consists of backbone amides or positively charged residues, stabilising the transition state lowers the activation energy necessary for the reaction, and so promotes catalysis. Additionally, it may allow for insertion or positioning of a substrate, enzymes that catalyse multi-step reactions can have multiple oxyanion holes that stabilise different transition states in the reaction. Enzyme catalysis Active site Transition state Serine proteases#Catalytic mechanism Albert Lehninger, et al
Protein Data Bank
The Protein Data Bank is a crystallographic database for the three-dimensional structural data of large biological molecules, such as proteins and nucleic acids. The PDB is overseen by a called the Worldwide Protein Data Bank. The PDB is a key resource in areas of structural biology, most major scientific journals, and some funding agencies, now require scientists to submit their structure data to the PDB. Many other databases use protein structures deposited in the PDB, for example, SCOP and CATH classify protein structures, while PDBsum provides a graphic overview of PDB entries using information from other sources, such as Gene ontology. By 1971, one of Meyers programs, SEARCH, enabled researchers to access information from the database to study protein structures offline. SEARCH was instrumental in enabling networking, thus marking the beginning of the PDB. Upon Hamiltons death in 1973, Tom Koeztle took over direction of the PDB for the subsequent 20 years, in January 1994, Joel Sussman of Israels Weizmann Institute of Science was appointed head of the PDB.
In October 1998, the PDB was transferred to the Research Collaboratory for Structural Bioinformatics, the new director was Helen M. Berman of Rutgers University. In 2003, with the formation of the wwPDB, the PDB became an international organization, the founding members are PDBe, RCSB, and PDBj. Each of the four members of wwPDB can act as deposition, data processing, the data processing refers to the fact that wwPDB staff review and annotate each submitted entry. The data are automatically checked for plausibility. The PDB database is updated weekly, the PDB holdings list is updated weekly. As of 14 March 2017, the breakdown of current holdings is as follows,103,514 structures in the PDB have a structure factor file,9,057 structures have an NMR restraint file. 2,826 structures in the PDB have a chemical shifts file, the final conformation of the protein is obtained, in the latter case, by solving a distance geometry problem. A few proteins are determined by cryo-electron microscopy, the significance of the structure factor files, mentioned above, is that, for PDB structures determined by X-ray diffraction that have a structure file, the electron density map may be viewed.
The data of such structures is stored on the electron density server, since 2007, the rate of accumulation of new protein structures appears to have plateaued. The file format used by the PDB was called the PDB file format. This original format was restricted by the width of computer punch cards to 80 characters per line, around 1996, the macromolecular Crystallographic Information file format, mmCIF, which is an extension of the CIF format started to be phased in
Cystathionine beta synthase
Cystathionine-β-synthase, known as CBS, is an enzyme that in humans is encoded by the CBS gene. This enzyme belongs to the family of lyases, to be specific, the hydro-lyases, CBS is a multidomain enzyme composed of an N-terminal enzymatic domain and two CBS domains. The CBS gene is the most common locus for mutations associated with homocystinuria, the systematic name of this enzyme class is L-serine hydro-lyase. Other names in use include, beta-thionase, cysteine synthase, L-serine hydro-lyase, methylcysteine synthase, serine sulfhydrase. Methylcysteine synthase was assigned the EC number EC126.96.36.199 in 1961, a side-reaction of CBS caused this. The EC number EC188.8.131.52 was deleted in 1972, the human enzyme cystathionine β-synthase is a tetramer and comprises 551 amino acids with a subunit molecular weight of 61 kDa. It displays a modular organization of three modules with the N-terminal heme domain followed by a core contains the PLP cofactor. The cofactor is deep in the domain and is linked by a Schiff base.
A Schiff base is a group containing a C=N bond with the nitrogen atom connected to an aryl or alkyl group. The heme domain is composed of 70 amino acids and it appears that the heme only exists in mammalian CBS and is absent in yeast and protozoan CBS. At the C-terminus, the domain of CBS contains a tandem repeat of two CBS domains of β-α-β-β-α, a secondary structure motif found in other proteins. CBS has a C-terminal inhibitory domain, the C-terminal domain of cystathionine β-synthase regulates its activity via both intrasteric and allosteric effects and is important for maintaining the tetrameric state of the protein. This inhibition is alleviated by binding of the effector, adoMet, or by deletion of the regulatory domain, however. Mutations in this domain are correlated with hereditary diseases, the heme domain contains an N-terminal loop that binds heme and provides the axial ligands C52 and H65. The presence of protoporphyrin IX in CBS is a unique PLP-dependent enzyme and is found in the mammalian CBS. D. melanogaster and D.
discoides have truncated N-terminal extensions, the Anopheles gambiae sequence has a longer N-terminal extension than the human enzyme and contains the conserved histidine and cysteine heme ligand residues like the human heme. The PLP is an internal aldimine and forms a Schiff base with K119 in the active site, between the catalytic and regulatory domains exists a hypersensitive site that causes proteolytic cleavage and produces a truncated dimeric enzyme that is more active than the original enzyme. Both truncated enzyme and the found in yeast are not regulated by adoMet
A cofactor is a non-protein chemical compound or metallic ion that is required for a proteins biological activity to happen. These proteins are enzymes, and cofactors can be considered helper molecules that assist in biochemical transformations. A coenzyme that is tightly or even covalently bound is termed a prosthetic group, the two subcategories under coenzyme are cosubstrates and prosthetic groups. Cosubstrates are transiently bound to the protein and will be released at some point, the prosthetic groups, on the other hand, are bound permanently to the protein. Both of them have the function, which is to facilitate the reaction of enzymes. Additionally, some sources limit the use of the cofactor to inorganic substances. An inactive enzyme without the cofactor is called an apoenzyme, while the enzyme with cofactor is called a holoenzyme. Some enzymes or enzyme complexes require several cofactors, organic cofactors are often vitamins or made from vitamins. Many contain the nucleotide adenosine monophosphate as part of their structures, such as ATP, coenzyme A, FAD and this common structure may reflect a common evolutionary origin as part of ribozymes in an ancient RNA world.
It has been suggested that the AMP part of the molecule can be considered to be a kind of handle by which the enzyme can grasp the coenzyme to switch it between different catalytic centers. Cofactors can be divided into two groups, organic cofactors, such as flavin or heme, and inorganic cofactors, such as the metal ions Mg2+, Cu+, Mn2+. Organic cofactors are sometimes divided into coenzymes and prosthetic groups. The term coenzyme refers specifically to enzymes and, as such, on the other hand, prosthetic group emphasizes the nature of the binding of a cofactor to a protein and, refers to a structural property. Different sources give different definitions of coenzymes, cofactors. It should be noted that terms are often used loosely. However, the author could not arrive at a single all-encompassing definition of a coenzyme, the study of these cofactors falls under the area of bioinorganic chemistry. In nutrition, the list of essential trace elements reflects their role as cofactors, in humans this list commonly includes iron, manganese, copper and molybdenum.
Although chromium deficiency causes impaired glucose tolerance, no human enzyme that uses this metal as a cofactor has been identified, iodine is an essential trace element, but this element is used as part of the structure of thyroid hormones rather than as an enzyme cofactor
Urocanase is the enzyme that catalyzes the second step in the degradation of histidine, the hydration of urocanate into imidazolonepropionate. Urocanase is coded for by the UROC1 gene, located on the 3rd chromosome in humans, the protein itself is composed of 676 amino acids which fold, producing the final product which has 2 identical subunits, making the enzyme a homodimer. To catalyze the hydrolysis of urocanate in the pathway of L-histidine the enzyme utilizes its two NAD+ groups. The NAD+ groups act as electrophiles, attaching to the top carbon of the urocanate which leads to sigmatropic rearrangement of the urocanate molecule and this rearrangement allows for the addition of a water molecule, converting the urocanate into 4, 5-dihydro-4-oxo-5-imidazolepropanoate. Urocanate + H2O ⇌4, 5-dihydro-4-oxo-5-imidazolepropanoate Inherited deficiency of urocanase leads to elevated levels of acid in the urine. Urocanase is found in bacteria, in the liver of many vertebrates and has been found in the plant Trifolium repens.
Urocanase is a protein of about 60 Kd, it binds tightly to NAD+, a conserved cysteine has been found to be important for the catalytic mechanism and could be involved in the binding of the NAD+. Urocanate Hydratase at the US National Library of Medicine Medical Subject Headings EC184.108.40.206
A chemical reaction is a process that leads to the transformation of one set of chemical substances to another. Nuclear chemistry is a sub-discipline of chemistry that involves the reactions of unstable. The substance initially involved in a reaction are called reactants or reagents. Chemical reactions are characterized by a chemical change, and they yield one or more products. Reactions often consist of a sequence of individual sub-steps, the elementary reactions. Chemical reactions are described with chemical equations, which present the starting materials, end products. Chemical reactions happen at a characteristic reaction rate at a given temperature, reaction rates increase with increasing temperature because there is more thermal energy available to reach the activation energy necessary for breaking bonds between atoms. Reactions may proceed in the forward or reverse direction until they go to completion or reach equilibrium, Reactions that proceed in the forward direction to approach equilibrium are often described as spontaneous, requiring no input of free energy to go forward.
Non-spontaneous reactions require input of energy to go forward. Different chemical reactions are used in combinations during chemical synthesis in order to obtain a desired product, in biochemistry, a consecutive series of chemical reactions form metabolic pathways. These reactions are catalyzed by protein enzymes. Chemical reactions such as combustion in fire and the reduction of ores to metals were known since antiquity, in the Middle Ages, chemical transformations were studied by Alchemists. They attempted, in particular, to lead into gold, for which purpose they used reactions of lead. The process involved heating of sulfate and nitrate minerals such as sulfate, alum. In the 17th century, Johann Rudolph Glauber produced hydrochloric acid and sodium sulfate by reacting sulfuric acid, further optimization of sulfuric acid technology resulted in the contact process in the 1880s, and the Haber process was developed in 1909–1910 for ammonia synthesis. From the 16th century, researchers including Jan Baptist van Helmont, Robert Boyle, the phlogiston theory was proposed in 1667 by Johann Joachim Becher.
It postulated the existence of an element called phlogiston, which was contained within combustible bodies. This proved to be false in 1785 by Antoine Lavoisier who found the explanation of the combustion as reaction with oxygen from the air
Enzyme promiscuity is the ability of an enzyme to catalyse a fortuitous side reaction in addition to its main reaction. Although enzymes are remarkably specific catalysts, they can often perform side reactions in addition to their main and these promiscuous activities are usually slow relative to the main activity and are under neutral selection. An example of this is the atrazine chlorohydrolase from Pseudomonas sp, ADP which evolved from melamine deaminase, which has very small promiscuous activity towards atrazine, a man-made chemical. Enzymes are evolved to catalyse a reaction on a particular substrate with a high catalytic efficiency. Several theoretical models exist to predict the order of duplication and specialisation events, on the other, enzymes may evolve an increased secondary activity with little loss to the primary activity with little adaptive conflict. A study of three distinct hydrolases has shown the main activity is robust towards change, whereas the activities are more plastic.
Specifically, selecting for an activity that is not the activity, does not initially diminish the main activity. The most recent and most clear cut example of evolution is the rise of bioremediating enzymes in the past 60 years. Due to the low number of amino acid changes, these provide an excellent model to investigate enzyme evolution in nature. This issue can be resolved thanks to ancestral reconstruction and this variability in ancestral specificity has not only been observed between different genes, but within the same gene family. Antithetically, the ancestor before the split had a more pronounced isomaltose-like glucosidase activity. Roy Jensen in 1976 theorised that primordial enzymes had to be highly promiscuous in order for networks to assemble in a patchwork fashion. This primordial catalytic versatility was lost in favour of highly catalytic specialised orthologous enzymes, as a consequence, many central-metabolic enzymes have structural homologues that diverged before the last universal common ancestor.
Promiscuity is however not only a primordial trait, in fact it is very widespread property in modern genomes, a series of experiments have been conducted to assess the distribution of promiscuous enzyme activities in E. coli. In E. coli 21 out of 104 single-gene knockouts tested could be rescued by overexpressing a noncognate E. coli protein, overexpressing the ORF collection allowed E. coli to gain over an order of magnitude in resistance in 86 out 237 toxic environment. Homologues are sometimes known to display promiscuity towards each others main reactions, despite the divergence the homologues have a varying degree of reciprocal promiscuity, the differences in promiscuity are due to mechanisms involved, particularly the intermediate required. Examples of these are enzymes for primary and secondary metabolism in plants, a promiscuous activity is a non-native activity the enzyme did not evolve to do, but arises due to an accommodating conformation of the active site. When the specificity of enzyme was probed, it was found that it was selective against natural amino acids that were not phenylalanine
Uroporphyrinogen III synthase
Uroporphyrinogen III synthase EC220.127.116.11 is an enzyme involved in the metabolism of the cyclic tetrapyrrole compound porphyrin. It is involved in the conversion of hydroxymethyl bilane into uroporphyrinogen III, the enzyme folds into two alpha/beta domains connected by a beta-ladder, the active site being located between the two domains. A deficiency is associated with Gunthers disease, known as congenital erythropoietic porphyria and this is an autosomal recessive inborn error of metabolism that results from the markedly deficient activity of uroporphyrinogen III synthase. Uroporphyrinogen III synthase at the US National Library of Medicine Medical Subject Headings This article incorporates text from the public domain Pfam and InterPro IPR003754
In biochemistry, allosteric regulation is the regulation of an enzyme by binding an effector molecule at a site other than the enzymes active site. The site to which the effector binds is termed the allosteric site, Allosteric sites allow effectors to bind to the protein, often resulting in a conformational change involving protein dynamics. Effectors that enhance the activity are referred to as allosteric activators. Allosteric regulations are an example of control loops, such as feedback from downstream products or feedforward from upstream substrates. Long-range allostery is especially important in cell signaling, Allosteric regulation is particularly important in the cells ability to adjust enzyme activity. The term allostery comes from the Greek allos and stereos and this is in reference to the fact that the regulatory site of an allosteric protein is physically distinct from its active site. Most allosteric effects can be explained by the concerted MWC model put forth by Monod and Changeux, or by the model described by Koshland, Nemethy.
Both postulate that enzyme subunits exist in one of two conformations, tensed or relaxed, and that relaxed subunits bind substrate more readily than those in the tense state, the two models differ most in their assumptions about subunit interaction and the preexistence of both states. Thus, all subunits must exist in the same conformation, the model further holds that, in the absence of any ligand, the equilibrium favors one of the conformational states, T or R. The equilibrium can be shifted to the R or T state through the binding of one ligand to a site that is different from the active site. The sequential model of allosteric regulation holds that subunits are not connected in such a way that a change in one induces a similar change in the others. Thus, all enzyme subunits do not necessitate the same conformation, the sequential model dictates that molecules of a substrate bind via an induced fit protocol. In general, when a subunit randomly collides with a molecule of substrate, while such an induced fit converts a subunit from the tensed state to relaxed state, it does not propagate the conformational change to adjacent subunits.
Instead, substrate-binding at one subunit only slightly alters the structure of other subunits so that their sites are more receptive to substrate. A morpheein is a structure that can exist as an ensemble of physiologically significant. Transitions between alternate morpheein assemblies involve oligomer dissociation, conformational change in the state, and reassembly to a different oligomer. The required oligomer disassembly step differentiates the morpheein model for allosteric regulation from the classic MWC, porphobilinogen synthase is the prototype morpheein. Ensemble models like the Ensemble Allosteric Model and Allosteric Ising Model assume that each domain of the system can adopt two states similar to the MWC model, molecular dynamics simulations can be used to estimate a systemss statistical ensemble so that it can be analyzed with the allostery landscape model
Enzyme catalysis is the increase in the rate of a chemical reaction by the active site of a protein. The protein catalyst may be part of a complex, and/or may transiently or permanently associate with a Cofactor. Catalysis of biochemical reactions in the cell is vital due to the very low rates of the uncatalysed reactions at room temperature and pressure. A key driver of protein evolution is the optimization of such catalytic activities via protein dynamics, the mechanism of enzyme catalysis is similar in principle to other types of chemical catalysis. By providing an alternative reaction route the enzyme reduces the required to reach the highest energy transition state of the reaction. The reduction of activation increases the amount of reactant molecules that achieve a sufficient level of energy, such that they reach the activation energy. As with other catalysts, the enzyme is not consumed during the reaction but is recycled such that a single enzyme performs many rounds of catalysis, the favored model for the enzyme-substrate interaction is the induced fit model.
The advantages of the induced fit mechanism arise due to the effect of strong enzyme binding. There are two different mechanisms of substrate binding, uniform binding, which has strong binding, and differential binding. The stabilizing effect of uniform binding increases both substrate and transition state binding affinity, while differential binding increases only transition state binding affinity, both are used by enzymes and have been evolutionarily chosen to minimize the activation energy of the reaction. It is important to clarify, that the induced fit concept cannot be used to rationalize catalysis and that is, the chemical catalysis is defined as the reduction of Ea‡ relative to Ea‡ in the uncatalyzed reaction in water. The induced fit only suggests that the barrier is lower in the form of the enzyme. Induced fit may be beneficial to the fidelity of molecular recognition in the presence of competition, →→→editor These conformational changes bring catalytic residues in the active site close to the chemical bonds in the substrate that will be altered in the reaction.
After binding takes place, one or more mechanisms of catalysis lowers the energy of the transition state. This effect is analogous to an increase in concentration of the reagents. The binding of the reagents to the enzyme gives the reaction intramolecular character, the situation might be more complex, since modern computational studies have established that traditional examples of proximity effects cannot be related directly to enzyme entropic effects. Also, the original proposal has been found to largely overestimate the contribution of orientation entropy to catalysis. Histidine is often the residue involved in these reactions, since it has a pKa close to neutral pH