ENZYME RELATED TO EXERCISE
ENZYME RELATED TO EXERCISE • Creatine kinase • Aminotransferases • Alanine aminotransferase (ALT) • Aspartate aminotransferase (AST) • Hexokinase, pyruvate kinase (glycolysis) • Phosphorylase, protein kinase A (glycogenolysis) • Hormone-sensitive lipase • Adenylate Cyclase • Lactate Dehydrogenase, Creatine Kinase (anaerobic system) • Pyruvate dehydrogenase • NADH dehydrogenase • Citrate synthase • ATP synthase • Acyl-coA dehydrogenase (aerobic system) • Transaminase (amino acid metabolism during exercise)
CREATINE • Creatine is a nitrogenous organic acid that exists naturally in most vertebrates. The role of creatine is to facilitate the recycling of adenosine diphosphate (ADP) to adenosine triphosphate (ATP) via the donation of phosphate groups. In tissues, creatine also acts as a pH buffer. • Creatine is synthesized in the liver and kidneys and can be obtained from diet. In human body, creatine and phosphocreatine are stored in skeletal muscle and distributed in blood, brain, and other tissues. • Creatine is transported through the blood and taken up by tissues with high energy demands, such as the brain and skeletal muscle, through an active transport system. The concentration of ATP in skeletal muscle is usually 2 – 5 mM, which would result in a muscle contraction of only a few seconds. During the times of increased energy demands, the phosphagen (or ATP/PCr) system rapidly resynthesizes ATP from ADP with the use of phosphocreatine (PCr) through a reversible reaction with the enzyme creatine kinase (CK).
CONTINUE… CREATINE KINASE • In skeletal muscle, the concentrations of PCr may reach 20 – 35 mM. In most muscles, the capacity of ATP regeneration of CK is very high and is therefore not a limiting factor. Although the cellular concentrations of ATP are small, changes are difficult to detect because ATP is continuously and efficiently replenished from the large pools of PCr and CK. Creatine has the ability to increase muscle stores of PCr thus potentially increasing the muscle’s ability to resynthesize ATP from ADP to meet increased energy demands. • Creatine kinase (CK) is also known as creatine phosphokinase (CPK) or phospho-creatine kinase. An enzyme expressed by various tissues and cell types, CK catalyzes the conversion of creatine and utilizes adenosine triphosphate (ATP) to create phosphocreatine (PCr) and adenosine diphosphate (ADP). This reaction is reversible, therefore, ATP can be generated from PCr and ADP.
ALANINE AMINOTRANSFERASE • Alanine transaminase (ALT) is a transaminase enzyme. It is also called alanine aminotransferase (ALAT) and was formerly called serum glutamate-pyruvate transaminase (SGPT) or serum glutamic-pyruvic transaminase (SGPT). ALT is found in plasma and various body tissues, and most common in the liver. It catalyzes the two parts of the alanine cycle: serum ALT level and serum AST (aspartate transaminase) level, and their ratio (AST/ALT ratio). These two parts of the alanine cycle are commonly measured clinically as biomarkers for liver health. • ALT catalyzes the transfer of an amino group from L-alanine to α-ketoglutarate, and the products of this reversible transamination reaction are pyruvate and L-glutamate, indicated as follows: L-alanine + α-ketoglutarate ⇌ pyruvate + L-glutama
ASPARTATE AMINOTRANSFERASE • Aspartate transaminase (AST) or aspartate aminotransferase is known as serum glutamic oxaloacetic transaminase (SGOT). AST catalyzes the reversible transfer of an α-amino group between aspartate and glutamate. AST is found in the liver, heart, skeletal muscle, kidneys, brain, and red blood cells. Serum AST level, serum ALT (alanine transaminase) level, and their ratio (AST/ALT ratio) are commonly measured clinically as biomarkers for liver health. The tests are part of blood panels. • Reaction is catalyzed by aspartate aminotransferase • As a prototypical transaminase, AST relies on PLP (Vitamin B6) as a cofactor to transfer the amino group from aspartate or glutamate to the corresponding ketoacid. In the process, the cofactor shuttles between PLP and the pyridoxamine phosphate (PMP) form. The transfer of the amino group, which is catalyzed by aspartate aminotransferase is crucial for the biosynthesis and degradation of amino acid.
PYRUVATE KINASE • Pyruvate kinase is an enzyme that catalyzes the final step of a glycolysis process. It catalyzes the transfer of a phosphate group from phosphoenolpyruvate (PEP) to adenosine diphosphate (ADP), yielding one molecule of pyruvate and one molecule of ATP. • Glycolysis • There are two steps in the reaction of pyruvate kinase in glycolysis. I.
First, PEP transfers a phosphate group to ADP - producing ATP and the enolate of pyruvate.
II. Secondly, a proton is added to the enolate of pyruvate to produce a functional form of pyruvate that the cell requires. Because the substrate for pyruvate kinase is a simple phospho-sugar and the product is an ATP. • The glycolytic reaction catalyzed by pyruvate kinase is the final step of glycolysis. This process is one of the three rate-affecting steps of the catabolic reaction cascade. The rate-affecting steps are the slower steps of a reaction and therefore determine the rate of the overall reaction. In glycolysis, the rate-affecting steps are coupled with the hydrolysis of ATP or the phosphorylation of ADP to create a highly energetically favorable and irreversible reaction mechanism. This final step is highly regulated and deliberately irreversible because pyruvate is a crucial intermediate building block for further metabolic pathways. Once pyruvate kinase synthesizes pyruvate, the pyruvate either enters the TCA cycle for further production of ATP under aerobic conditions or is reduced to lactate under anaerobic conditions.
CONTINUE
• Gluconeogenesis: the reverse reaction • Pyruvate kinase also serves as a regulatory enzyme for gluconeogenesis, which is a biochemical pathway in which the liver generates glucose from pyruvate and other substrates. Gluconeogenesis utilizes noncarbohydrate sources to provide glucose to the brain and red blood cells in times of starvation when direct glucose reserves are exhausted.
PROTEIN KINASE
• Protein kinase is a kinase enzyme that modifies other proteins by chemically adding phosphate groups to them by a process known as phosphorylation. • Phosphorylation usually results in a functional change of the target protein (substrate) by changing enzyme activity and cellular location, or by association with other proteins. • The chemical activity of a kinase involves transferring a phosphate group from a nucleoside triphosphate (usually ATP) and covalently attaching it to specific amino acids with a free hydroxyl group.
HORMONE-SENSITIVE LIPASE • Hormone-sensitive lipase is previously known as cholesteryl ester hydrolase (CEH). The main function of hormone-sensitive lipase is to mobilize stored fats. • Mobilization and Cellular Uptake of Stored Fats • HSL functions to hydrolyze either a fatty acid from a triacylglycerol molecule, thus freeing the fatty acid and diglyceride, or the fatty acid from a diacylglycerol molecule, thus freeing the fatty acid and monoglyceride. • HSL is activated when the body needs to mobilize energy stores, and hence responds positively to catecholamine, ACTH. HSL is inhibited by insulin. • Another important role is the release of cholesterol from cholesterol esters for use in the production of steroids.
ADENYLATE CYCLASE • Adenylyl cyclase is commonly known as adenyl cyclase or adenylate cyclase ( AC). • AC performs key regulatory roles in all cells. Six distinct classes of AC have been described, all catalyzing the same reaction but representing unrelated gene families with no known sequence or structural homology. • The best known class of adenylyl cyclases is class III or AC-III (Roman numerals are used for classes). ACIII occurs widely in eukaryotes and has important roles in many human tissues. • All classes of adenylyl cyclases catalyze the conversion of adenosine triphosphate (ATP) to 3',5'-cyclic AMP (cAMP) and pyrophosphate. Magnesium ions are generally required and appear to be closely involved in the enzymatic mechanism.
LACTATE DEHYDROGENASE (ANAEROBIC SYSTEM) • Lactate dehydrogenase (LDH or LD) is an enzyme found in nearly all living cells (animals, plants, and prokaryotes). LDH catalyzes the conversion of lactate to pyruvic acid and back, as it converts NAD+ to NADH and back. A dehydrogenase is an enzyme that transfers a hydride from one molecule to another. • LDH is expressed extensively in body tissues, such as blood cells and heart muscle. Because it is released during tissue damage, it is the marker of common injuries and diseases, including heart failure.
ROLE IN MUSCULAR FATIGUE • The onset of acidosis during the periods of intense exercise is commonly attributed to the accumulation of lactic acid. • From this reasoning, the idea of lactate production being a primary cause of muscle fatigue during exercise has been widely adopted. A closer mechanistic analysis of lactate production under anaerobic conditions shows no biochemical evidence of the production of lactate through LDH, which can contribute to acidosis. • LDH works to prevent muscular failure and fatigue in multiple ways. The lactate-forming reaction generates cytosolic NAD+, which feeds into the glyceraldehyde 3-phosphate dehydrogenase reaction to help maintain cytosolic redox potential and promote substrate flux through the second phase of glycolysis, which in turn, promotes ATP generation. This outcome, in effect, provides more energy to the contracting muscles under heavy workloads. The production and removal of lactate from cells also eject a proton consumed in the LDH reaction. The removal of excess protons produced in the wake of this fermentation reaction serves as a buffer system for muscle acidosis. Once the proton accumulation exceeds the rate of uptake in lactate production and removal through the LDH symport, muscular acidosis occurs.
PYRUVATE DEHYDROGENASE
• Pyruvate dehydrogenase contributes to transforming pyruvate into acetyl-CoA by a process called pyruvate decarboxylation. • Acetyl-CoA may then be used in the citric acid cycle to carry out cellular respiration; therefore, pyruvate dehydrogenase contributes to linking the glycolysis metabolic pathway to the citric acid cycle and to releasing energy via NADH.
NADH DEHYDROGENASE
• NADH dehydrogenase is an enzyme with a systematic name called NADH: acceptor oxidoreductase. This enzyme catalyzes the following chemical reaction. • NADH dehydrogenase is a flavoprotein that contains iron-sulfur centres. NADH dehydrogenase is used in the electron transport chain for the generation of ATP.
CITRATE SYNTHASE • The enzyme citrate synthase exists in nearly all living cells and stands as a pace-making enzyme in the first step of the citric acid cycle. Citrate synthase is localized within eukaryotic cells in the mitochondrial matrix. • Citrate synthase is commonly used as a quantitative enzyme marker for the presence of intact mitochondria. The maximal activity of citrate synthase indicates the mitochondrial content of skeletal muscle, and the activity can be increased by endurance training or high-intensity interval training. • Citrate synthase catalyzes the condensation reaction of two-carbon acetate residue from acetyl coenzyme A and a molecule of four-carbon oxaloacetate to form the six-carbon citrate, as indicated below: • acetyl-CoA + oxaloacetate + H2O → citrate + CoA-SH • Oxaloacetate is regenerated after the completion of one round of the Krebs cycle. Oxaloacetate is the first substrate to bind to the enzyme, and the binding induces the enzyme to change its conformation and creates a binding site for the acetylCoA. Only when this citroyl-CoA has formed will another conformational change cause thioester hydrolysis and release coenzyme A. This outcome ensures that the energy released from the thioester bond cleavage drive the condensation.
CONTINUE… INHIBITION
• Citrate synthase is inhibited by high ratios of ATP:ADP, acetyl-CoA:CoA, and NADH:NAD because high concentrations of ATP, acetyl-CoA, and NADH show that the energy supply is high for the cell. • It is also inhibited by succinyl-CoA, which resembles Acetyl-coA and acts as an uncompetitive inhibitor.
ATP SYNTHASE • ATP synthase is an enzyme that creates the energy storage molecule adenosine triphosphate (ATP). For most organisms, ATP is the most commonly used "energy currency" of cells. ATP is formed from adenosine diphosphate (ADP) and inorganic phosphate (Pi). The overall reaction catalyzed by ATP synthase is ADP + Pi + H+out ⇌ ATP + H2O + H+in • The formation of ATP from ADP and Pi is energetically unfavorable and would normally proceed in reverse direction. In order to drive this reaction forward, ATPase couples ATP synthesis (during cellular respiration) with an electrochemical gradient created by the difference in proton (H+) concentration across the mitochondrial membrane in eukaryotes or the plasma membrane in bacteria. During photosynthesis in plants, ATP is synthesized by ATPase using proton gradient created in the thylakoid lumen through the thylakoid membrane and into the chloroplast stroma.
ACETYL COENZYME A DEHYDROGENASE • Acyl-CoA dehydrogenases (ACADs) belong to a class of enzymes that functions to catalyze the initial step in each cycle of fatty acid β-oxidation in the mitochondria of cells. The action of ACADs results in the introduction of a trans double-bond between C2 (α) and C3 (β) of the acyl-CoA thioester substrate. • ACADs can be categorized into three distinct groups based on their specificity for short-, medium-, or long-chain fatty acid acyl-CoA substrates. Although different dehydrogenases target fatty acids of varying chain length, all types of ACADs are mechanistically similar. Differences in the enzyme are due to the location of the active site along the amino acid sequence. • ACADs constitute an important class of enzymes in mammalian cells due to their role in metabolizing the fatty acids in ingested food materials. The action of this enzyme represents the first step of fatty acid metabolism, which is the process of breaking long chains of fatty acids into acetyl CoA molecules.
TRANSAMINASE • Transaminases or aminotransferases are enzymes that catalyze a transamination reaction between an amino acid and an α-keto acid. These enzymes are important for the synthesis of amino acids, which form proteins. • An amino acid contains an amine (NH2) group. A keto acid contains a keto (=O) group. In transamination, the NH2 group on one molecule is exchanged with the =O group on the other molecule. The amino acid becomes a keto acid, and the keto acid becomes an amino acid. • Many transamination reactions occur in tissues and are catalyzed by the transaminases specific for a particular amino/keto acid pair. These reactions are readily reversible, and the direction is determined by the reactants that are in excess.
ENZYME RELATED TO RECOVERY
• Glycogen synthase (glycogenesis) • Fatty acid synthase (fatty acid synthesis)
GLYCOGEN SYNTHASE
• Glycogen synthase is a key enzyme in glycogenesis, which is the conversion of glucose to glycogen. This enzyme is a glycosyltransferase, and glycogen is an enzyme that combines excess glucose residues one by one into a polymeric chain for storage as glycogen. • Glycogen synthase concentration is highest in the bloodstream 30 to 60 minutes after an intense exercise.
FATTY ACID SYNTHASE • In humans, fatty acid synthase (FAS) is an enzyme encoded by the FASN gene. Fatty acid synthase is a multi-enzyme protein that catalyzes fatty acid synthesis. It is not a single enzyme but a whole enzymatic system composed of two identical 272 kDa multifunctional polypeptides, in which substrates are handed from one functional domain to the next. • The main function of fatty acid synthase is to catalyze the synthesis of palmitate (a long-chain saturated fatty acid) from acetyl-CoA and malonyl-CoA in the presence of NADPH. • Fatty acids are aliphatic acids that are fundamental to cellular structure and to the production and storage of energy. Fatty acids also act as intermediates in the biosynthesis of hormones and other biologically important molecules.
EFFECTS OF EXERCISE ON ENZYME REGULATION • Exercise speed up glycolysis in muscle – increase glycolysis enzymes activity • Exercise speed up pyruvate oxidation in muscle – increase activity of pyruvate dehydrogenase • Exercise speed up citric acid cycle in muscle – increase CAC enzymes activity • Exercise speed up oxidative phosphorylation in muscle – increase ETC enzymes activity • Exercise speed up lipolysis – increase lipase activity in adipocyte and muscle • Exercise speed up fatty acid oxidation in muscle – increase ß-oxidation enzymes activity