BIOCHEMISTRY (I) OF HUMAN ENZYMES AND METABOLISM

The elections develop the material described above.

The tutorials (in two groups of students, 1-2 teachers per group) summarize and deepen the material using examples of clinical cases (problem-based learning) that highlight the application of biochemical knowledge in the diagnosis and treatment of serious diseases. In each tutorial, the students prepare the answers to the questions related to the examined clinical cases so that they can discuss them with the teachers.

The laboratory exercises (in 4 groups of students, 3 teachers per group of 30 students) constitute the students' practical training, are a necessary supplement to the lectures and aim to familiarize them with the use of techniques, the operation of instruments and the conduct of tests that are often used in Clinical Biochemistry, as well as helping to understand concepts that are not easily understood theoretically (learning based on practical experience).

Attendance is mandatory in all tutorials and laboratory exercises.

Information and Communication Technologies are used to prepare lecture materials and provide online information and learning aids to students.

Specifically:

• Common software (eg powerpoint) is used to prepare lecture material and display slides and videos.
• The study guide (explanatory material & additional bibliography), the tutorial material (clinical cases for processing), the theory and protocols of the laboratory exercises, the handover slides after each lesson as well as videos and scientific articles related to the taught material are made electronically and online available to students through the e-class.
• Information about the course, the teachers and the

their research interests and the Biochemistry Laboratory in general are available online on the Laboratory's website http://www.med.uth.gr/biochemistry/index.html

• Common software (eg excel) is used for the statistical processing of student evaluation
• Announcements, information, etc. are available online through e-class. Communication is also done via e-mail.

The assessment language of the students is Greek.

Evaluation methods.

A. In the laboratory exercises: Laboratory Work, Reference, Written Exams with short answer and problem solving questions.

The students' participation in the exercises as well the written report of the results of the exercises, which is done by the students during the workshops, is mandatory. The report includes the measurements (presented in tables and plots), and the conclusions (eg whether the expected results were obtained, if not why not, sources of possible errors in the experiments) as requested by each exercise. At the end of each exercise, the written report is checked by the teachers and signed if it has been filled in correctly. In the event that particularly demanding calculations or diagrams need to be made, it is possible for the report to be completed after the end of the workshop and delivered to the lecturers at a later predetermined date. Successful participation in the exercise is certified by the teacher's signature on the written report of the results.

At the end of the semester, students are examined in writing on the content of the Workshops. On Laboratory exams material to be examined is the theory, the methodology and the methods of processing results included in the Laboratory Exercises Guide or developed by the lecturers during the laboratories. Only those who successfully participated in the laboratory exercises have the right to participate in the Laboratory exams. Passing the laboratory exams is a necessary condition for participation in the course exams.

B. In the tutorials:  Public Presentation, Oral Examination.

In each tutorial, the students prepare the answers to the questions related to the examined clinical cases so that they can present them and discuss them with the lecturers. The material of the tutorials is examined in writing together with the material of the lectures.

C. In theory – lecture material: Written Exams with multiple-choice and short-answer questions.

The course exams are written, lasting 1.5 hours, and consist of multiple-choice questions (examples are available in the Course Guide) and critical or short-answer questions related to clinical cases depending on those discussed in the tutorials. Examinable material is the material of lectures and tutorials as described above. Only those students who have passed the Laboratory exams have the right to participate in the course exams.

Final grade

The final grade of the course is calculated by 80% from the grade of the written exams of the course and by 20% from the grade of the written exams of the Laboratory.

All of the above can be accessed by the students as they are contained in the Course Guide which is distributed in print to all students and is posted electronically in the e-class.

The general purpose of the course is to introduce students to the understanding of the normal and pathological functioning of the human organism at the molecular and cellular level and to the principles and techniques of laboratory medicine. Also, to enable them to design, analyze and apply the knowledge of biochemistry in clinical practice, ie the diagnosis and treatment of diseases.

Furthermore, the course seeks to provide students with a foundation upon which the student will build to support the knowledge provided in the courses of longer semesters and specifically in the courses: "Biochemistry of Gene Expression, Human Organs and Functions", "Systems Physiology" , "Neurophysiology & Endocrine Physiology", "Clinical Biochemistry", "Pharmacology", "Pathological Physiology", "Medical Genetics", "Pathology" and "Pediatrics".

Finally, the aim of the course is for the students to understand the importance of the correct execution and assessment of laboratory analyzes and the corresponding required skills for both Laboratory Medicine specialties and for the clinical specialties of Medicine.

The specific objectives of the course are specialized in the following intended learning outcomes:

Upon successful completion of the course, the student:

He/she will be able to use the acquired knowledge in order to:

• describes the basic metabolic functions and the molecular-biochemical basis of the metabolic diseases of the human body.
• describes the structure and properties of proteins and enzymes, the ways of production, consumption and storage of energy and the intermediate metabolism of the main biomolecules and the mechanisms of metabolic regulation.
• has the resources to distinguish symptoms and suggest diagnosis and treatment of basic metabolic diseases.
• make use of the basic equipment of a biochemical/clinical chemistry laboratory and carry out basic biochemical analyses.
• analyze and process the results of basic biochemical analyses
• collaborate with fellow students in a laboratory environment to perform basic biochemical analyzes and process their results.

General Skills

• Search, analysis and synthesis of data and information, using the necessary technologies
• Adaptation to new situations
• Decision making
• Autonomous work
• Teamwork
• Exercise criticism and self-criticism
• Promotion of free, creative and inductive thinking

A. Theoretical clinical skills
I. By theory chapter
01. Introduction to Biochemistry:
• Biochemical tests as a means of diagnosis
• Diagnosis & treatment of metabolic diseases
• Biochemical basis of drug action
02. Enzymes: Basic Properties, Kinetics, Regulation & Catalytic Mechanisms
• Protease inhibitors as antihypertensive & HIV drugs
• Carbonic anhydrase inhibitors as anticancer drugs
• Applications of restriction nucleases in genetic testing
• Lactate dehydrogenase isoenzymes in clinical diagnosis
03. Oxygen transport proteins (hemoglobin) and connective tissue (collagen)
Biochemical basis, diagnosis and treatment
• sickle cell disease
• thalassemia
• scurvy
• Ehlers Danlos syndromes
• incomplete osteogenesis
04. Introduction to metabolism,
• Niacin deficiency
05. Digestion and degradation of carbohydrates, glycolysis
Biochemical basis, diagnosis and treatment
• pyruvate kinase deficiency
• lactic acidosis
• galactosemia
• fructose intolerance
• lactose intolerance
06. Gluconeogenesis and regulation of blood glucose
• Structure and action of insulin
• Structure and action of glucagon
• Biochemical basis and treatment of hypoglycemia
07. Citric acid cycle
Biochemical basis, diagnosis and treatment
• pyruvate dehydrogenase phosphatase deficiency
• beri-beri disease
• mercury or arsenic poisoning
08. Respiratory chain – Oxidative phosphorylation – ATP synthesis
• Pathological action & treatment of free oxygen radicals (ROS)
• Biochemical basis of cyanide poisoning
• Gray adipose tissue and thermoregulation
• Biochemical basis of mitochondrial diseases
09. Pentoses phosphate pathway
• Biochemical basis, diagnosis and treatment of G6PD deficiency
• Thiamine deficiency
• Wernicke-Korsakoff syndrome
• Principles of mass neonatal screening
10. Glycogen metabolism & glucose homeostasis
Biochemical basis, diagnosis and treatment
• glycogen storage diseases
• Pombe disease
• Von Gierke's disease
• McArdle's disease
• hypoglycemia
11. Lipid digestion, fatty acid degradation, ketone bodies
Biochemical basis, diagnosis and treatment
• carnitine deficiency
• Vitamin deficiency. B12 – megaloblastic anemia
• Zellweger syndrome
• diabetic ketoacidosis
12. Biosynthesis and regulation of lipid metabolism
• Nutritional causes & treatment of obesity
• Structure, metabolism and action of non-steroidal anti-inflammatory drugs
13. Membrane lipids and structure of biological membranes
Biochemical basis, diagnosis and treatment
• sphingolipids
• Tay-Sachs disease
14. Metabolism of cholesterol and lipoproteins
• Structure, metabolism and action of bile salts
• Biochemical basis of cholelithiasis
• Structure, metabolism and action of vitamin D
• Biochemical basis of childhood rickets
• Biochemical basis, diagnosis and treatment of hyperlipidemias
• Familial hypercholesterolemia
• Biochemical basis of atherosclerosis
15. Digestion and degradation of proteins and amino acids, urea cycle
• Clinical application of transaminases
• Biochemical basis, diagnosis and treatment of hyperammonemia
16. Biosynthesis of amino acids, heme metabolism
Biochemical basis, diagnosis and treatment
• albinism
• phenylketonuria
• hyperhomocysteinemia
• porphyria
• hyperbilirubinemia
17. Nucleotide metabolism
• Lesch-Nyhan syndrome
• Biochemical action of cytotoxic anticancer drugs
• Biochemical basis, diagnosis and treatment of gout
18. Summary and completion of metabolism
• Effect of starvation on health
• Biochemical basis, diagnosis and treatment of diabetes mellitus
• Effect of alcohol on health
19. Metabolic diseases and metabolic regulation
• Definition and categories of metabolic diseases
• Introduction to the diagnosis, management and treatment of hereditary metabolic diseases.
20. Metabolism & cancer
• Properties of cancer cells
• Principles of PET application in cancer diagnosis
• Targeted anticancer drugs
II. In compulsory schools – problem-based learning
Presentation, analysis, discussion and treatment of real clinical cases
patients with the following pathological conditions:
• Penicillin resistance
• Incomplete osteogenesis
• Sickle cell disease
• Antitrypsin deficiency – emphysema,
• Alcoholism
• G6PD deficiency – drug-induced hemolytic anemia
• Pyruvate dehydrogenase deficiency
• Fructose intolerance
• Glycogen storage disease I
• Anorexia nervosa
• Obesity – Non-alcoholic steatohepatitis
• Hyperhomocysteinemia – Myocardial infarction
• Hyperammonemia
• Acetyl-CoA carboxylase deficiency.

B. Practical and clinical skills
In the mandatory workshops – learning based on practical experience
Acquisition of general and specific practical skills by the medical students during the preparation
of laboratory exercises:
1. General skills
• observation of phenomena in the laboratory, recording and analysis of data
• drawing conclusions from data
• solving practical problems
• building graphs from data
• extracting information from graphs
• handling basic equipment of Clinical Chemistry and Biochemistry laboratories
• effective team work
• safe work in a laboratory environment
• following and following instructions
• perception of laboratory errors and identification of their sources
2. Special skills
Clinical Applications of Enzymes and Enzyme Activity Analysis:
• The loss of enzyme activity as a cause of genetic diseases (enzymopathies)
• Determination of enzyme activity photometrically
• Enzymes as drugs
• Enzymes as diagnostic indicators of pathological conditions (enzyme diagnosis)
• Determination of enzyme concentration photometrically
• Diagnostic significance of alkaline phosphatase determination
• Enzymes as tools for the determination of metabolites
• Effect of substrate on enzyme reaction rate
• Determination of enzyme kinetic constants
• Enzyme inhibitors as poisons and drugs
• Enzyme inhibitor characterization
• Evaluation of laboratory results
Clinical Application of Hemoglobin Laboratory Analysis:
• Pathological causes of decrease or increase of hemoglobin in the blood
• Determination of hemoglobin concentration in the blood of patients photometrically
• Hemoglobin diseases
• Mutated hemoglobins
• Detection of abnormal hemoglobins by electrophoresis
• Evaluation of laboratory results
Clinical application of laboratory analysis of blood lipids
• Classification and characteristics of hyperlipidemias
• Cardiovascular diseases
• Determination of triglycerides in patient sera photometrically
• Measurement of total, HDL and LDL cholesterol in patient sera.
• Evaluation of laboratory results

C. Knowledge – Lecture material

Introduction to Biochemistry
• Biochemistry as a science: Definition, object, history
• Biochemistry in everyday life
• Biochemistry as basic knowledge for Medicine:
o Biopathology specialty
o Laboratory diagnosis, "Biochemical analyzes or tests"
o Basis of hereditary and metabolic diseases
o Understanding drug action and discovering new therapeutic substances
o Using and targeting metabolism to treat cancer
• Biochemistry as a course: Writings, prerequisite knowledge, organization, schedule, obligations

Enzymes: Basic Properties, Kinetics, Regulation & Catalytic Mechanisms
(Repeat/summary of Medicinal Chemistry chapters)
• Basic properties and working principles of enzymes, kinetics.
• Basic principles of enzyme activity regulation
• Catalysis Basics & Enzyme Inhibitors
• Serine protease catalysis strategies

Enzyme applications in diagnosis & treatment
• Enzymes as diagnostic markers
• Isoenzymes (lactic dehydrogenase, alkaline phosphatase, creatine kinase) in
diagnosis
• Enzymes as diagnostic tools: Applications of restriction endonucleases in genetic testing
• Enzymes and enzyme inhibitors as drugs: carbonic anhydrase inhibitors, protease inhibitors.
• Allosteric enzymes as new drug targets

Oxygen binding & transport proteins – Myoglobin & Hemoglobin
• general structure of heme
• the characteristics and main amino acids of the heme pocket
• myoglobin O2 binding curve
• structure and subgroups of hemoglobin & O2 binding curve
• molecular basis and importance of the allosteric effect of hemoglobin
• effect of 2,3-BPG and the BOHR effect
• molecular basis and detection of sickle cell anemia, thalassemias.

Connective tissue proteins – Collagen
• the components of connective tissue
• structure, characteristic composition, modifications, biosynthesis and role of collagen
• structure, characteristic composition, modifications and role of elastin
• structure, characteristic composition and role of proteoglycans and glycosaminoglycans
• genetic and pathological abnormalities of the connective tissue (scurvy, lathyrism, Ehlers-Danlos syndromes, osteogenesis imperfecta)
• Clinical example: Vitamin C deficiency

Introduction to metabolism
• Basic concepts of metabolism, relationship between anabolism and catabolism, common patterns of reactions
• Free energy of reaction and its importance for the concept of coupling reactions
• Role of ATP and ways of its production
• Role of redox coenzymes (NAD, niacin, FAD, NADP) and acetyl-coenzyme A.
• Niacin deficiency
• Role of creatine phosphate in muscles
• Ways of regulating metabolic processes and energy load of the cell.

Regulation of metabolism
• Regulation of metabolism through cell signal transduction
• Basic pathways of signal transduction – molecular circuits
• Structure and function of seven transmembrane helix receptors, G proteins and adenylate cyclase
• Role of cyclic AMP as an intracellular messenger and importance of the "reaction cascade"

Carbohydrate digestion
• Biological role and chemical nature of glucose and other basic carbohydrates in the diet
• Reactions and enzymes of carbohydrate digestion
• GLUT glucose transporters

Glycolysis
• Reactions of glycolysis and the enzymes that catalyze them
• Energy efficiency of glycolysis
• Metabolic fates of pyruvate and maintenance of redox balance
• Importance of glycolysis as an anaerobic pathway
• Pyruvate kinase deficiency
• Lactate management by the human body, lactic acidosis

Hormonal regulation of glycolysis & Gluconeogenesis
• General regulation of glycolysis, action of insulin and glucagon
• Hormonal regulation of phosphofructokinase and role of 2,6, fructose diphosphate
• Importance of isozymes for tissue metabolic specificity (hexokinase/glucokinase)
• Regulation during muscle rest and intense muscle exercise
• Gluconeogenesis reactions and energy costs
• Regulation of irreversible reactions
• Balanced regulation of glycolysis-gluconeogenesis
• Cori's cycle and Alanine cycle.

Metabolism of galactose & fructose
• Galactose metabolism
• Galactosemia
• Lactose intolerance
• Metabolism of fructose
• Fructose intolerance

The citric acid cycle
• Conversion of pyruvate to acetyl-coenzyme A in the mitochondria
• Krebs cycle reactions and the enzymes that catalyze them
• Substitute reactions of the cycle
• Cycle intermediates that are biosynthetic precursors
• Pyruvate dehydrogenase phosphatase deficiency
• Biochemical basis of beri-beri disease and mercury or arsenic poisoning

Regulation of the citric acid cycle
• Regulation of pyruvate dehydrogenase reaction
• Regulation of key enzymes and energy efficiency of cycle reactions
• Cycle regulation by energy load and hormones
• Clinical example: Pyruvate carboxylase deficiency

Oxidative Phosphorylation: Respiratory Chain
• Complexes and mobile electron transporters of the respiratory chain
• Electron transfer prosthetic groups, its structure and evolution
cytochrome c
• Generation of oxygen radicals (ROS) and combating them
• Chemosmotic effect (Mitchel hypothesis).
• Modes of transport of cytoplasmic NADH electrons to the mitochondrion
(glycerol phosphate and malate/aspartate systems)

Oxidative Phosphorylation: Synthesis of ATP
• General structure and function of F0F1-ATPase
• Proton motive force and coupling of the respiratory chain to oxidative phosphorylation
• Transport of ATP in and out of the mitochondrion.
• Energy efficiency of the respiratory chain and the complete burning of glucose
• Regulation of oxidative phosphorylation (respiratory control).

Thermogenesis & mitochondrial diseases
• Inhibition and uncoupling of oxidative phosphorylation
• Thermogenesis and gray adipose tissue
• Mitochondrial diseases
• Role of mitochondria in apoptosis, cancer and aging

Glycogen Metabolism
• Structure, physical and chemical properties of glycogen.
• Degradation of glycogen and regulation of glycogen phosphorylase
allosteric and hormonal
• Glycogen synthesis, glycogenin and regulation of glycogen synthase.
• Importance of glycogenolysis and glycogenogenesis in the liver and muscles
• Role of protein phosphatase 1 (PP1) in the regulation of glycogen metabolism
& phosphorylase a as a glucose sensing system in the liver

Glucose homeostasis & glycogen storage diseases
• Role of glycogen in glucose homeostasis
• The effect of glucagon and insulin on glycogen metabolism
• Glycogen storage diseases (general and specific to Pombe, Von Gierke and McArdle diseases)

Pentose phosphate pathway
• Importance of the pentose phosphate pathway in NADPH and pentose production and its coordination with glycolysis
• In which webs it is active, when and why
• General oxidative and non-oxidative branch reactions.
• Implications and biochemical basis of G6PD deficiency (drug-induced hemolytic
anemia, favism)
• Thiamine deficiency and Wernicke-Korsakoff syndrome

Lipid digestion & fatty acid degradation
• Structural characteristics and nomenclature of lipids
• Comparison of energy storage efficiency in the form of triacylglycerols or
glycogen
• Digestion of food lipids: organs, enzymes, chylomicrons
• Mobilization of lipids from adipose tissue: hormone-sensitive lipases
• Utilization of glycerol of lipids
• Activation of fatty acids and their transport to the mitochondrion (role of carnitine)
• β-oxidation of fatty acids and energy efficiency
• Degradation of fatty acids with an odd number of carbon atoms
• Importance and role of vitamin B12 (cobalamin)
• Carnitine deficiency
• Vitamin deficiency. B12 – megaloblastic anemia

Composition & role of ketone bodies
• Degradation of fatty acids in the absence of glucose
• Ketone bodies
o production
the role
the metabolism
o importance in fasting and diabetes
o diabetic ketoacidosis

Fatty acid synthesis, regulation & dietetics
• Basic reactions and energy costs of fatty acid biosynthesis (differences with degradation)
• Importance and origin of acetyl-CoA and NADPH required
• Regulation and coordination of fatty acid degradation and synthesis in response to nutrition by acetyl-CoA carboxylase
• Classic and ketogenic weight loss diets
• Essential fatty acids,
• Cyclooxygenase and biochemical basis of action of non-steroidal anti-inflammatory drugs

Composition of storage lipids, cholesterol & derivatives
• Synthesis of triacylglycerols,
• Biological roles of cholesterol & basic principles of its synthesis
• HMG-CoA reductase and its regulation
• Structure and function of bile salts, cholelithiasis
• Synthesis of steroid hormones – Cytochrome P450
• Composition and role of vitamin D, childhood rickets

Membrane lipids & membrane structure
• Structure, biological role and composition of phospholipids, sphingolipids and gangliosides,
• Sphingolipidoses, respiratory distress syndrome and Tay-Sachs disease
• Structural characteristics, chemical composition and properties of biological membranes
• Factors affecting the fluidity of membranes

Lipoproteins & atherosclerosis
• Classification, composition, function and diagnostic significance of lipoproteins
• Structure and function of the LDL receptor and its importance in hypercholesterolemia and atherosclerosis
• Familial hypercholesterolemia
• Biochemical basis of atherosclerosis
• Biochemical basis of pharmaceutical treatment of hypercholesterolemia
• Clinical example: Hypertriglyceridemia – lipoprotein lipase deficiency

Digestion and degradation of proteins & amino acids
• Process of digestion of food proteins
• Process of intracellular degradation of proteins
• General principles and tissue specificity of amino acid catabolism
• Nitrogen transport and release reactions: transamination, oxidative and hydrolytic deamination, glutamine hydrolysis.
• Importance and role of vitamin B6 and pyridoxal phosphate
• Clinical significance of transaminases

Nitrogen catabolism & urea cycle
• Role of alanine and glutamine in the transport of amino groups in the liver
• General principles of nitrogen excretion
• Reactions of the urea cycle
• Molecular basis, importance and treatment of hyperammonemias

Amino acid metabolism
• General principles of catabolism of the carbon skeleton of amino acids
• Glycogenetic and ketogenic amino acids
• Genetic diseases of amino acid catabolism, albinism
• Molecular basis, diagnosis and treatment of phenylketonuria
• Main pathways of nitrogen incorporation into biomolecules
• Essential and non-essential amino acids
• Synthesis reactions of non-essential amino acids

Methyl Cycle & Amino Acid Derivatives
• Transfer reactions of monocarbonate groups
• Role of tetrahydrofolic acid and S-adenosylmethionine
• Normal and pathological role of homocysteine, hyperhomocysteinemia
• Biologically important derivatives of amino acids: biogenic amines, catecholamines, glutathione, nitric oxide

Heme & iron metabolism
• General principles of heme biosynthesis and catabolism
• Biological significance and excretion of bilirubin
• Hyperbilirubinemia, Jaundice
• Molecular basis and symptoms of the porphyrias
• Biological functions of iron
• The daily cycle of iron
• Abnormalities in iron metabolism
• Maintenance of iron homeostasis

Nucleotide metabolism
• Structure, nomenclature and biological role of nucleotides
• Precursor compounds and basic principles for the de novo synthesis of ribonucleotides & excess pathways
• Biochemical basis and symptoms of Lesch-Nyhan disease
• Synthesis of deoxyribonucleotides from ribonucleotides
• Regulation of the synthesis of nucleotides
• Products of nucleotide catabolism and biochemical basis of gout.
• Anticancer drugs that target tetrahydrofolate reductase and thymidylate synthase

Summary & integration of metabolism
• Repetitive patterns in metabolic regulation
• Main metabolic processes, connections and their control points
• Important node molecules: glucose 6-phosphate, pyruvate and acetyl-CoA
• Main metabolic processes of brain, muscle (skeletal and cardiac), adipose tissue, kidney and liver
• Main hormones of metabolism
• Metabolic changes at the organismal level during the postprandial and fasting state
• Metabolic disorders in diabetes mellitus
• Metabolic changes during exercise and alcohol consumption

Summary of Metabolic Diseases
• Definition and categories of metabolic diseases
• Biochemical basis of clinical symptoms
• Diagnosis, treatment and treatment of hereditary metabolic diseases.

Metabolism & Cancer
• The special characteristics of cancer cells
• The Warburg effect (aerobic glycolysis)
• Importance of the Warburg effect in the diagnosis and treatment of cancer
• Metabolic oncogenes and tumor suppressors, tumor metabolites
• Hypoxia and cancer
• Hypoxia-inducible HIF factors and their role in cancer
Modern research and targeting of cancer metabolism