Cellular Biology & Homeostasis Enzymes - Part 3 Ross University PDF

Cellular Biology & Homeostasis ENZYMES - Part 3 VP 2025 Clara Camargo, DVM, Cert AqV  LEARNING OBJECTIVES 1. Explain cofactors (coenzyme, cosubstrate, prosthetic group) and give some examples 2. Understand what is a zymogen and its physiological relevance, give examples 3....

Cellular Biology & Homeostasis ENZYMES - Part 3 VP 2025 Clara Camargo, DVM, Cert AqV  LEARNING OBJECTIVES 1. Explain cofactors (coenzyme, cosubstrate, prosthetic group) and give some examples 2. Understand what is a zymogen and its physiological relevance, give examples 3. Define apoptosis and generally explain how it works 4. Describe the mechanisms of enzyme regulation COFACTORS The catalytic activity of many enzymes Cofactor Enzyme Organic molecules depends on the presence of small Thiamine pyrophosphate Pyruvate dehydrogenase Flavin adenine dinucleotide (FADH) Monoamine oxidase molecules called: cofactors Nicotinamide adenine dinucleotide (NADH) Lactate dehydrogenase  small non-protein molecules Pyridoxal phosphate Glycogen phosphorylase Coenzyme A (CoA) Acetyl CoA carboxylase  “helpers” Biotin Pyruvate carboxylase 5′-Deoxyadenosyl cobalamin Methylmalonyl mutase Tetrahydrofolate (THF) Thymidylate synthase Metal Cofactors can be subdivided into two 2+ Zn Carbonic anhydrase 2+ Zn Carboxypeptidase groups: Mg 2+ EcoRV 2+ Mg Hexokinase 1. Inorganic metals Ni 2+ Urease Mo Nitrate reductase 2. Small organic molecules (coenzymes) Se Glutathione peroxidase 2+ Mn Superoxide dismutase + K Propionyl CoA carboxylase COFACTORS  Cofactors Enzymes can exist in inactive forms (apoenzyme) and be converted to active forms (holoenzyme) → with the help of a cofactor  If the cofactor is a small organic molecule  it is called coenzyme Apoenzyme → Coenzyme binding → Holoenzyme  Tightly bound (covalent bonds) → prosthetic groups (Heme group)  Loosely bond → co-substrates bind to and are released from the enzyme just as substrates and products are. (NADPH) Many enzymes acquire full enzymatic activity after they are fully folded (recall the post-translational modifications) and bound to the cofactor/coenzyme. Fe bind to the heme group in hemoglobin COENZYMES Coenzymes are often derived from vitamins Can be either tightly or loosely bound to the enzyme Can be associated with the enzyme’s active site that assists with their catalytic function  Vitamins cannot be FYI synthesized by humans and most animals; and must be supplied from the diet.  Many vitamins are essential components of enzymes or provide those as coenzymes COENZYMES - Biotin Attached to distinct lysine residues in histones, affecting chromatin structure and mediating gene regulation Recall (epigenetic modification) ZYMOGENS  ZYMOGENS Some enzymes are synthesized as inactive precursors that are activated by proteolytic cleavage of one or a few specific peptide bonds. The inactive precursor is called a zymogen (or a proenzyme) The biochemical change usually occurs in the Golgi apparatus, or when digestive enzymes are secreted in the organ lumen (i.e., duodenum) ENZYME ACTIVATION BY PROTEOLYSIS Specific proteolysis is common in cellular physiology  The digestive enzymes are synthesized as zymogens in the stomach and pancreas. I.e.: o Pepsinogen → pepsin (stomach) o Trypsinogen → Trypsin (pancreas)  The pancreas and stomach secretes zymogens partly to prevent the enzymes from digesting the cells in which they are synthesized Not only are digestive enzymes synthesized Accidental activation of zymogens can as inactive precursors, but many protein happen when the pancreatic secretion duct hormones are also produced in this form is blocked resulting in acute pancreatitis i.e.; preproinsulin → proinsulin → insulin SECRETION OF ZYMOGEN GRANULES BY CELLS OF THE PANCREAS Zymogens Exocrine pancreas cells have many small granules of zymogens that are visible microscopically Darker-staining cells form clusters called acini, which are arranged in lobes separated by a thin fibrous barrier The secretory cells of each acinus surround a small intercalated duct  ZYMOGENS Proteolytic cleavage = activation For the cleavage, energy (ATP) is not required. Proteolytic cleavage occurs just once in the life of an enzyme molecule, the process is irreversible. Unlike allosteric control and reversible covalent modification  DIETARY PROTEIN DIGESTION BY PROTEASES - Stomach Site of Proenzyme/ Active enzyme  Initially, pepsinogens are converted to pepsin synthesis Zymogen Stomach Pepsinogen Pepsin in the gastric lumen by gastric acid (HCl) Pancreas Chymotrypsinogen Chymotrypsin  Once this reaction begins, pepsin can Pancreas Trypsinogen Trypsin Pancreas Procarboxypeptidase Carboxypeptidase autocatalyze the conversion of pepsinogen to Pancreas Proelastase Elastase pepsin steps to regulate enzyme activation Gastric gland DIETARY PROTEIN DIGESTION BY PROTEASES - Pancreas Site of Proenzyme/ Active enzyme synthesis Zymogen Stomach Pepsinogen Pepsin zymOGEN or PROenzyme in the pancreas Pancreas Chymotrypsinogen Chymotrypsin Pancreas Trypsinogen Trypsin Pancreas Procarboxypeptidase Carboxypeptidase Pancreas Proelastase Elastase Cascade of events regulating digestive enzymes Stomach Proteins Proenteropeptidase Enteropeptidase Pepsin Trypsinogen Trypsin Large peptides Chymotrypsinogen Chymotrypsin Gastric and pancreatic Proelastase Elastase Procarboxypeptidases A and B Carboxypeptidases A and B zymogens Pancreatic prolipase Pancreatic lipase Small peptides Aminopeptidases Dipeptidases Tripeptidases Free amino acids + Triglycerides ZYMOGEN AND COENZYME - COAGULATION CASCADE Blood clotting is mediated by a cascade of FYI proteolytic activations that ensures a rapid and amplified response to trauma. VITAMIN K (Coenzyme) → coenzyme participating in the synthesis of blood coagulation factors II, VII, IX and X FYI WARFARIN (drug which reduces blood clot formation) → Compete irreversibly with a liver enzyme (epoxy reductase complex) → Depletion of active Vit K ZYMOGEN - APOPTOSIS Programmed cell death, or apoptosis, is mediated by proteolytic enzymes called caspases, which are synthesized as zymogens (procaspases) within the cells. When activated, caspases function to cause cell death in most multicellular organisms. It produces special cell fragments (apoptotic bodies), which are cleared by macrophages. Unlike necrosis (traumatic cell death), apoptosis is highly regulated and does not cause inflammation ZYMOGEN - APOPTOSIS Initiator caspases  executioner caspases kill the cell by degrading proteins indiscriminately Cannot stop once it has begun FYI Can be initiated through: intrinsic pathway extrinsic pathway  Both pathways use caspases (proteases) Supplemental video – necrosis and apoptosis Necrosis vs Apoptosis https://www.youtube.com/watch?v=zFrBwGfOQs0&t=246s  OTHER ROLES OF ZYMOGENS IN BIOLOGY Zymogens Many developmental processes are controlled by the activation of zymogens. For example, in the metamorphosis of a tadpole into a frog, large amounts of collagen are resorbed from the tail. The conversion of procollagenase into collagenase (the active protease) is precisely timed in these From: Alberts. Mol. Biol. of the Cell remodeling processes.  Likewise, much collagen is broken down in a mammalian uterus after delivery.  ENZYME REGULATION Enzyme inhibition and regulation both affect enzyme activity, but they differ in their mechanisms and outcomes: Enzyme inhibition typically involves a specific molecule binding to an enzyme, reducing its activity and preventing substrate conversion  This can be reversible or irreversible In contrast, enzyme regulation encompasses a broader range of processes that modulate enzyme activity, often in response to cellular conditions  Regulation can involve allosteric sites, covalent modifications, or feedback mechanisms, allowing for fine-tuned control over metabolic pathways While inhibition often leads to a decrease in activity, regulation can enhance or diminish enzyme function as needed. MECHANISMS FOR REGULATING ENZYME ACTIVITY An organism must coordinate its different metabolic processes by:  Regulating the reaction velocity of enzymes depending on the substrate concentration (Km range) (increased substrate → increased reaction rate) Some enzymes with specialized regulatory functions can be regulated when physiologic conditions change, by changing: Enzyme activity Gene expression (Induction and repression of enzyme synthesis/degradation) MECHANISMS FOR REGULATING ENZYME ACTIVITY REGULATION OF ENZYME ACTIVITY ALLOSTERIC ENZYMES  ↑ or ↓ affinity for substrate  Modify Vmax Allosteric enzymes change shape upon binding of an effector Effectors (modifiers/regulators) bind noncovalently at a site other than the active site  Altering the affinity of the enzyme for its substrate OR  Modifying the maximal catalytic activity of the enzyme https://www.youtube.com/watch?v=ApKM-IkSElY REGULATION OF ENZYME ACTIVITY ALLOSTERIC ENZYMES Positive effector Negative effector Modify maximal catalytic velocity (Vmax) Effectors can influence affinity of enzyme for its substrate (K0.5) Both  REGULATION OF ENZYME ACTIVITY ALLOSTERIC ENZYMES Homotropic effectors: when the substrate itself serves as an effector Most allosteric substrates function as positive homotropic effectors: the presence of the substrate molecule at one site of the enzyme enhances the catalytic properties of the other substrate-binding From: Harvey. Biochemistry sites Hemoglobin is a homotropic allosteric protein REGULATION OF ENZYME ACTIVITY ALLOSTERIC ENZYMES Heterotropic effectors: effector different from the substrate Recall FA synthesis Classical example is a feedback inhibition of a metabolic pathway (end-product inhibition)  REGULATION OF ENZYME ACTIVITY COVALENT MODIFICATION (Reversible) Covalent modifications: usually addition or removal of phosphate groups from specific amino acids of the enzyme (Ser, Tyr, Thr). Phosphorylation reactions are catalyzed by kinases using ATP as a phosphate donor. The phosphorylated protein may be more or less active. i.e., glycogen metabolism REGULATION OF ENZYME ACTIVITY ENZYME SYNTHESIS Enzymes regulated by synthesis are typically those required only at specific stages of development or under certain physiological conditions.  This allows the organism to conserve resources and respond effectively to changing needs Can you give some examples? Induction or repression of protein synthesis are slow (hours to days), compared with allosteric or covalent regulation of enzyme activity Why? REGULATION OF ENZYME ACTIVITY - Summary Regulation of allosteric enzymes Covalent modification of enzymes Induction or repression of enzymes synthesis HAPPY STUDYING [email protected] ©2021 Ross University School of Veterinary Medicine. All rights reserved.