coenzyme--1.pdf

30 Ola Qutaiba Malak Salameh Nafez Abu-Tarboush Enzyme-Cofactors During catalysis, most enzymes rely on their active sites, on the other hand some might get the help from small molecules, in order to get through the reaction, these molecules can be classified into Organic molecules or non-organic...

30 Ola Qutaiba Malak Salameh Nafez Abu-Tarboush Enzyme-Cofactors During catalysis, most enzymes rely on their active sites, on the other hand some might get the help from small molecules, in order to get through the reaction, these molecules can be classified into Organic molecules or non-organic ones (metals), they get along with the enzyme in the reaction either to facilitate or participate in it, and so we call these molecules as (COFACTORS). [During Catalysis], some enzymes rely on the A) Amino Acids’ functional group (ex: Chymotyrpsin): -Almost all polar amino acids participate in nucleophilic catalysis. -Ser, Cys, Lys, & His can participate in covalent catalysis. -Histidine: because its Pka is near the physiological pH & acid–base catalysis. Note: The normal single histidine’s pka=6 but when it is a part of the polypeptide its PH is near the physiological PH is around 6.8~7.4. B) some rely on Coenzymes in Catalysis: - Usually (but not always) synthesised from vitamins. - Each coenzyme is specific for a type of reaction. - They are either: 1-Activation-transfer coenzymes. (activation of covalent bond with substrate then breaking it) 2-Oxidation–reduction coenzymes. -Note: (coenzymes are organic cofactors so, all coenzymes are cofactors but not all cofactors are coenzymes. For example, metal ions are cofactors but not coenzymes). VITAMINS are organic substances > we have 13 type, 4 of them are lipid soluble the rest are water soluble. Coenzymes: >modified vitamins< -When metals are tightly bound to proteins (covalently bonded), the proteins are called Metalloproteins, however when they are loosely bound (by non-covalent bonds, electrostatic interactions mainly); the proteins are called Metalo-associated protein. -When organic molecules are tightly bound to the protein (covalent bond), the co-enzymes are called prosthetic groups, however when they are loosely bound (non-covalent bonds); they are called co-substrates. Activation-Transfer Coenzymes: The functional group of the coenzyme directly participates in catalysis by Covalent bonding, they participate in the reaction, not facilitators, they themselves move groups. -Characteristics: Two groups in the coenzyme’s structure: -A functional group that forms a covalent bond with substrate, participate in the reaction. -A binding group that binds tightly to the enzyme, it binds to the same enzyme and becomes part of the active site. 2 Dependence on the enzyme for additional specificity of substrate & additional catalytic power. 1-Thiamine Pyrophosphate (TPP): The source is THIAMINE (B1). It is involved in decarboxylation reactions. (The molecule will lose 1 carbon in form of CO 2). Well, you can clearly see the 2 negatively charged oxygen atoms in the pyrophosphate compound, that is helping actually in stabilising the active site, because that is the binding site of the coenzyme. Reactive thiamine carbon forms a covalent bond with a substrate keto group while cleaving the adjacent carboncarbon bond (terminal one). What is the purpose of thiamin pyrophosphate? The pyrophosphate provides negatively charged oxygen atoms and chelates Mg+2 that is tightly bound to the enzyme, 2 oxygen atoms linked to Mg+2 and the Mg+2 is covalently linked with the enzyme, so the pyrophosphate is a part of the active site of the enzyme, linked via Mg+2. α-ketoglutarate dehydrogenase,Decarboxylation of α-ketoglutarate into succinyl CoA by αketoglutarate dehydrogenase, α-ketoglutarate dehydrogenase (α-KGDH) is an enzyme complex, similar to the pyruvate dehydrogenase complex. 3 Binding group is pyrophosphate, the functional group is the carbon that falls between the sulfur and nitrogen, this carbon will link to the pyruvate then there will be electron rearrangement, where the pyruvate loses CO 2, Decarboxylation of pyruvate into acetyl CoA is one of the important reactions catalysed by Pyruvate dehydrogenase complex & α-ketoglutarate dehydrogenase. 2- Coenzyme A (CoA): Source: pantothenate (B5): made of al βalanine and pantoic acid. Functional group: sulfhydryl group (nucleophile), Attacks carbonyl groups & forms acyl thioesters (the “A”). The bond between the carbon and the sulfur is HIGH ENERGY BOND. At the end of glycolysis, the conversion of pyruvate to acetyl CoA takes place, enabling its entry into the Krebs cycle. This transformation is facilitated by the pyruvate dehydrogenase complex, involving the assistance of both TPP and CoA. Following this, within the mitochondria, acetyl CoA combines with oxaloacetate to give citrate by the enzyme citrate synthase, a process that is aided by CoA. 4 3- Biotin (B7): Biotin is required for carboxylation reactions (covalently bound to Lys). You can find it in Food & intestinal bacteria. Deficiencies are seen after long antibiotic therapies (because that kills the intestinal bacteria) or excessive consumption of raw eggs (egg white protein, avidin, has a high affinity for biotin). Egg white is referred to as avidin which binds to biotin making it unavailable for working as a co-factor. There are many enzymes that use Biotin like: 1- Pyruvate carboxylase By adding the CO2 to the Pyruvate, we will get the oxaloacetate. 2-Acetyl CoA carboxylase (fatty acid synthesis) Here the Acetyl CoA is carboxylated into Malonyl CoA, in order to synthesis lipids. 5 4- Pyridoxal Phosphate (PLP): Sources: pyridoxal, pyridoxamine and pyridoxine. (Vitamin B6). Metabolism of amino acids via reversible transamination reactions. Upon the entry of an amino acid into the active site, the amino group forms a covalent linkage with the aldehyde present. As a result, an electron rearrangement occurs, leading to the departure of the amino acid from the active site as a ketoacid, devoid of its amino group. Simultaneously, the amino group becomes attached to a vitamin molecule. Following this, a distinct ketoacid (distinct from the one generated in the initial reaction) binds to the active site. It undergoes a reverse reaction within the site, leading to its exit as an amino acid. 6 Oxidation-Reduction Coenzymes: A large number of Coenzymes, they DON’T bind to the substrate, but ratherly bind to the Enzyme itself. Most common: NAD+ (niacin, B3) & FAD (riboflavin, B2) Others: work with metals to transfer single electrons to O2 (Vitamins E & C) Again: Dependence on the enzyme for additional specificity of substrate & additional catalytic power. 1- NAD+ (nicotinamide adenine dinucleotide): The functional group (C opposite to N) accepts a hydride ion from the substrate, dissociates, & a keto group (CO) is formed, the nitrogen in the ring helps to increase the electrophilicity. In the context of NAD+ and NADP+, the niacin ring functions as the reactive group, while the nucleotide segment of the molecule serves as the binding group that engages and attaches to the enzyme. At carbon C2, an R group is present. If this R group is a hydrogen (H), the resulting molecule is NAD+. Conversely, if the R group takes the form of a phosphate group, the resulting molecule is NADP+. The phosphate doesn’t participate in the reaction, but it rather determines which enzyme will NAD+ or NADP+ bind to. 7 The substrate undergoes oxidation, initiating with the detachment of a hydrogen atom from the substrate, which then associates with the ring structure as a hydride ion. Following this, the second hydrogen atom dissociates as a hydrogen ion, leading to the creation of a double bond within the substrate as a keto group, exemplified in our case by the conversion of lactate into pyruvate. NAD+ is usually involved in catabolism reactions while NADP+ is involved in anabolism reactions, this specificity is due to the presence or absence of the phosphate group. The enzyme’s histidine binds the proton of (-OH) on lactate making it easier for NAD+ to pull off the other hydrogen with both electrons (a hydride). A keto group (-C=O) is formed. 2- FAD & FMN: Both are prosthetic groups of flavoproteins. FMN = Flavin Mononucleotide. FAD = Flavin Adenine Dinucleotide. FAD is composed of FMN and an adenine nucleotide. FAD accepts electrons in a sequential manner, resulting in a time gap between the acceptance of the first and second hydrogen atoms. Consequently, when the first hydrogen atom attaches, FAD generates a radical referred to as FADH+. This radical is highly reactive and possesses potential harm to the cell. To prevent detrimental reactions with other hydrogen atoms, FAD becomes covalently bonded to the enzyme, which restricts its ability to damage the cell. It patiently awaits the binding of the next hydrogen atom after the formation of FADH2 takes place, eliminating the presence of the damaging radicals. 8 The source is RiboFlavin (B2) Succinate dehydrogenase Oxidation of succinate into fumarate by succinate dehydrogenase. Succinate dehydrogenase catalyzes the oxidation of succinate to fumarate. During this process, FAD (Flavin Adenine Dinucleotide) is reduced to FADH2. A Double bond is formed in the product. Also, the Pyruvate dehydrogenate complex involve in that type of reaction. ‫قراءة وفي منهم رح نرجع‬ ‫نوخدهم الفصل الجاي‬ 9 - Catalytic Metals: ‫احفظوه كامل رح ييجي عليه سؤال مباشر‬ Phosphofructokinase & TPP; (Mg2+) is required to coordinate the phosphate groups on the ATP for a successful reaction (chelation). Fructose-6-phosphate + ATP → fructose-1,6-bisphosphate + ADP. Some metals do not participate in enzyme catalysis directly but facilitate a reaction, like in the enzyme “alcohol dehydrogenase”, this reaction involves the reduction of NAD+ and the oxidation of the alcohol. If you look at the figure, ethanol (the alcohol) doesn’t bind to either the active site of the enzyme or to NAD+, so how does it bind? Histidine pulls the electrons from serine which in turn pulls the electrons from ethanol oxidizing it into an acetaldehyde. Then those electrons will be donated to NAD+ in the form of hydride ion. During those intermediates, the charges are stabilized by the zinc. So, even though the zinc doesn’t participate in the reaction, it facilitates it by stabilizing the charges. The histidine of alcohol dehydrogenase pulls a proton off the active site’s serine. The serine pulls off the proton of the substrate’s OH- group, leaving the oxygen with a negative charge. The charge is stabilized by zinc. A hydride is then transferred to NAD+. 10 Metalloenzymes - Metal ions are usually incorporated during synthesis & removal of the metal causes denaturation. -These metal ions may contribute either to the structure or the catalytic mechanism. - Liver alcohol dehydrogenase (dimer); 2 Zn+2 in each monomer; one for structural maintenance (joins the two subunits), the other is catalytic. - Carbonic anhydrase; A zinc atom is essentially always bound to four or more groups. 11