Biochemistry

Introduction to Biochemistry

06/10/13

BIOCHEMISTRY Biochemistry is the chemical basis of life Aim molecular level understanding of cell chemical processes from the previous goal, we can understand the origins of life as well as maintain its health Biochemistry directly relates to other branches of science such as physiology, immunology, toxicology, and etc. As future physicians, biochemistry offers us: a foundational understanding for our career continual learning acquisition of current knowledge emphasize the immensity of the unknown

Complete physical, mental, and social well being and not merely the absence of disease and infirmity, - WHO From a biochemical viewpoint, health is the optimal rate of intra- and extracellular reactions that is essential to the organism's maximal survival in the physiologic state Thus, biochemistry has a major impact in our nutritionand preventive medicine because it provides us understanding at the molecular level The Human Genome Project, completed in 2003 has provided us new insights on how the human body operates at the molecular level. Also started the boom on other fields such as proteomics, transcriptomics, glycomics, and etc.

less toxic form (removing H ion), leaving a low Km/affinity for acetyldehyde Developed only Cystolic high Km form which leads to high steady-state level of acetyldehyde in the blood resulting to increase sensitivity to alcohol and eventually flushing of the face Aspirin as treatment of heart disease Cycloxygenase: enzyme responsible for producing Prostaglandin (mediates sensation of pain) and Thromboxane (responsible for producing platelet aggregation for blood clotting resulting to heart attack) Aspirin blocks enzymatic action of Cycloxygenenase which inhibits Thromboxane production resulting to lesser chance of heart attacks but produces further bleeding in return. No platelet aggregation -> No clot formation -> No obstruction of coronary vessel -> NO HEART ATTACK! Role of Carnitine in fat metabolism To lose weight -> lose fats in our adipose tissue (made up of triacyglycerol, a glycerol esterified with fatty) Carnitine functions to transport fatty acid to the mitochondrion where oxidation (burning) of fatty acids takes place Fat transformed into energy instead of being stored Effects of Glutathione on skin Glutathione synthesizes Pheomelanin which produces reddish-brown skin pigment which results to whiter complexion Cell Types: 1. Eukaryotes Presence of membrane-enclosed nucleus encapsulation the DNA 2. Prokaryotes Lack discrete nucleus and no nuclear membrane (reason for fast cell replication)

Human Body 64% Water 20% Protein 13% Carbohydrates 10% Fat 5% Minerals BIOCHEMISTRY: Chemistry exploring Carbon atoms Catenation: Unique property of Carbon Ability to bond to its identical atom Biological applications of Biochemistry Facial flushing after drinking beer Asians: Genetic defect of mitochondrial Aldehyde Hydrogenase - an enzyme responsible for breaking down alcohol to

Human Cell Organelles Centrioles: for pulling chromatids Lysosomes: contain digestive enzymes and perform Endocytosis to digest of recycle foreign materials inside the cell (Suicide Sacks) Mitochondria: produce energy for the cell in the form of ATP and has their own DNA (Powerhouse) Nucleolus: Contains the genetic material of the cell and it is where ribosomes are assembled Nucleoplasm: where DNA floats

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Introduction to Biochemistry

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Rough Endoplasmic Reticulum: contains plenty of ribosomes and is responsible for protein synthesis Smooth Endoplasmic Reticulum: responsible for synthesis of fats and steroids Golgi Apparatus: performs quality control mechanism and glycosylation (addition of sugar) Peroxisomes: contain oxidative enzymes (catalase) for metabolism of fats Cytoplasm or Cytosol: medium where organelles can be found Plasma membrane: surrounds the cell and is responsible for communicating to other cells Cytoskeleton: maintains cell shape, responsible for cell movement, and arrangement and internal motion of cell organelles Microtubules (constituent of cilia) Microfilaments (Actin and Myosin)

mass spectrometry, use of enzymes, use of acid/alkaline hydrolysis, sequencing techniques, etc. 3. Analysis of Biomolecule function and metabolism Studies done on different levels such as whole animal, isolated parts, etc.

WATER AND PH I. Water Water ability to dissolve wide range of molecules dipole structure attraction of hydrogen electrons by oxygen gives it a partial positive charge 2 unshared electron pairs gives it a partial negative charge high capacity of h-bond formation regulated by hypothalamic mechanisms plays a role in pH due to its slight propensity to dissociate into hydroxide ions and protons tetrahedral in conformation with 2 3 hydrogens and 2 sp -hybridized orbitals as its corners can act both as an acid or base high dielectric constant (water = 78.5) greatly decreases the force of attraction between charged particles due waters high dielectric constant excellent nucleophile nucleophile e rich but does not necessarily posses a negative or positive charge H2O has 2 lone pairs of partially negative e making it an excellent nucleophile Hydrolysis - nucleophilic attack by water resulting to the cleavage of amide, glycoside or ester bonds Enzymes accelerate the rate of hydrolytic reactions Proteases catalyze the hydrolysis of proteins into amino acids Nucleases catalyze the hydrolysis of the phosphoester bonds in DNA and RNA

Plant Cell Cell wall: contains cellulose and is responsible for cellular strength Chloroplast: where mitochondria is located Vacuole: storage site Subcellular Fractionation: Process of isolating specific organelle 1. Extraction extreme pH, ostmotic pressure and temperature should be avoided 2. Homogenation disrupts the cell to liberate its constituents and suspends substances 3. Centrifugation specimen is subjected to greater speed, each yielding a pellet and supernatant (ex. nuclear fraction, mitochondrial fraction, microsomal fraction, etc.) ORGANELLES Nucleus Mitochondria Endoplasmic Reticulum Lysosomes Plasma membrane Golgi Apparatus Peroxisomes Cytosol MARKER DNA Glutamic dehydrogenase Glucose 6-Phosphatase Acid Phosphalase + + Na -K ATPase Galactosyltransferase Catalase, Uric Acid Oxidase Lactate dehydrogenase

Experimental approach used in Biochemistry 1. Isolation of Biomolecule and Organelles Ex. salt fractionation, chromatography, gel electrophoresis, etc. 2. Determination of Biomolecule structure Ex. elemental analysis, UV visible infrared, NMR spectroscopy, X-ray, crystallography,

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Introduction to Biochemistry

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A. Biomedical importance Dissolve most organic and inorganic molecules because of its dipolar structure and ability to form hydrogen bonds Excellent nucleophile 50-60% in adults 75% in children 60% intracellular ; 40% extracellular Tetrahedral structure: O2 in the center Dipole: electronegativity of water High dielectric constant: aka permittivity wherein it is a measure of the ability of a material to resist formation of electrical field (Separation force) B. Hydrolysis Water is capable of breaking down bonds C. Dissociation of water Water can act both as an acid and as a base ability of water to ionize. + Additional H = protonated form + H is removed = unprotonated Dissociation Constant (Ka) pKa = -log Ka o As the hydrogen dissociates, it becomes + more basic. At high pH, the H is already dissociated. o The stronger the acid, the lower is its pKa value (inversely proportional). o Dissociation will only occur completely in strong acid/ base Lower temp = Kw Higher temp = Kw o For pure water at 25 C = pH 7.0 + Low pH denotes high H pH power of hydrogen the negative log of the hydrogen ion concentration pH = -log [H ] the value of pH is inversely proportional + to the concentrations of H
+

Acid proton donors Base proton acceptors Strong acids complete dissociation even in strongly acidic solutions (low pH) Weak acids partial dissociation in acidic solutions Strong bases completely dissociates even strongly basic solutions (high pH) Dissociation constant (Ka) used to calculate the concentration of [H+], since weak electrolytes dissociates only partially
+

Acids protonated (HA or R-NH3 ) Conjugate Bases unprotonated (A or R-NH2) Weak Acid R-CH2-COOH Conjugate Base R-CH2-COO
-

* When H is remove from a weak acid it becomes a conjugate base. pKa = -logKa stronger the acid the lower itspKa value

The relative strengths of bases are expressed in terms of the pKa of their conjugate acids.

pKa = pH The pKa of an acid group is the pH at which the protonated and unprotonated species are present at equal concentrations.

Covalent Bond

Strongest force that holds the molecules together - Relatively weak and transient - influences H2Os high viscosity, high surface tension, high boiling point Self-association of non polar compounds in an aqueous

Hydrogen Bond

Hydrophobic

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Introduction to Biochemistry

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Interactions Electrostatic Interactions

environment Interactions between a oppositely charged groups Salt Bridges - Attraction between transient dipoles due to rapid movement of e of all neutral atoms - Significantly weaker than Hbonds

Tx: higher intake of water or give other base such as Bicarbonate

Alkalosis profuse vomiting excretesHCl from stomach (result: acid) o food poisoning o Tx: control vomiting, through medications and then the kidney will try to compensate

Van der Waals Forces

Diabetes Insipidus D. Henderson-Hasselbalch Equation describes the behavior of weak acids and buffers pH = pKa + log [A ]/ [HA] Where: pH = measure of acidity pKa = constant [A ] = Concentration of dissociated acid [HA] = concentration of the whole solution Carbonic anhydrase produces carbonic acid through the dissociation of CO2 and H2O and will form bicarbonate as an effective buffer system pKa = -log{[A-] [H+]} E. Buffer Systems neutralize any change of pH in your body used to resist pH changes by virtue of weak acids/bases and its conjugate 3 Important Physiologic Buffer Systems 1) Bicarbonate 2) Inorganic Orthophosphate 3) Intracellular Proteins such as hemoglobin Examples: Lungs Acidosis: accumulation of Lactic Acid Lactic acid = breathing Deeper breathing releases more CO2 to release excess acid. Kidneys protein = acid Excess protein is released as urine, which is acidic. Diabetic ketoacidosis Lack of insulin production No ADH urine output BIOLOGICAL MEMBRANES I. Membrane Separate inside from outside of a cell Separate inside from outside of an organelle But also enables selective material exchange Localize enzymes and enzyme functions Functions in excitation-response coupling Provides sites for energy transduction Compartmentalizes body water! Functions in excitation response coupling (act as receptor) To communicate with external environment Passing energy from one to the other o Are amphipatic (polar and nonpolarportion) o Hydrophobic (apolar tail) and hydrophilic (polar head) o Have saturated and/or unsaturated fatty acids as tails o More unsaturated (have kinks) = more fluid o More saturated (straight) = more rigid

Structure and Function Overview of Plasma Membrane highly fluid and dynamic lateral diffusion transverse movement (flip-flop) Fluid Mosaic Model dependent on lipid composition higher unsaturation = higher permeability longer fatty acid chain = lower permeability

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Introduction to Biochemistry

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bilayer made of lipids due to its amphipathic nature selective permeability outside-inside asymmetry outside choline-containing phospholipids inside aminophospholipids also contains proteins and carbohydrates

Cell organelles are also bounded by membranes localizes enzymes excitation-response coupling energy transduction changes may affect water balance and ion influx 60-40-20 rule 60% of lean body mass is water 40% is ICF 20% is ECF 5% is Plasma 15% is Interstitial ICF specialized environment for the cell energy utilization repair replicated cell functions ECF delivery system can be considered a highway for nutrients, hormones, and other substances Composition Substances ECF ICF NA K
+ 2+ 2+ +

II. Lipids A. Phospholipids Phosphoglycerides (more common) Sphingomyelin (prominent in myelin sheets B. Glycosphingolipids they have sugar components on the exterior of the cell C. Sterols Hydroxyl group is the polar portion cholesterol Cholesterol is the most common membranes of animal cells D. Micelle E. Lipid bilayer

of in

III. Membrane proteins Proteins o integral proteins interacts directly to the hydrophobic core of the bilayer usually globular needs detergent to solubize Peripheral proteins does not interact directly to the hydrophobic core bound to hydrophilic regions of integral proteins does not need detergent o Major functional molecules of membranes Enzymes Transporters Pumps and channels Structural components Antigens Receptors TABLE ENZYMATIC MARKERS Enzymatic Markers of Different Membranes Membrane Enzyme Plasma 5-nucleotidase Adenylyl cyclase + + Na -K -ATPase Endoplasmic reticulum Gluscose-6-phosphatase Golgi apparatus GlcNactransferase I Cis Goligimannosidase II Medial Galactosyltransferase Trans Sialyltrnasferase TGN Inner mitochondrial ATP synthase membrane o

140 mmol/L 4 mmol/L 2.5 mmol/L 1.5 mmol/L 100 mmol/L

-

10 mmol/L 140 mmol/L 0.1 umol/L 30 mmol/L 4 mmol/L 10 mmol/L 60 mmol/L 0-1 mmol/L 16 g/dL

CA CL

MG
-

HCO3
3PO4

27 mmol/L 2 mmol/L 5.5 mmol/L 2 g/dL

Glucose Protein

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Biochemistry
Introduction to Biochemistry

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Artificial Membranes used as models for membrane function varied lipid content choice of membrane proteins controlled environment liposomes for entrapment Lipid Rafts involved in signal transduction enriched in cholesterol, sphingolipids, and proteins Caveolae contains caveolin-1 derived from lipid rafts flask-shaped signal transduction Tight Junctions below the apical surface of epithelial cells prevents diffusion of macromolecules composed of proteins Other specializations desmosomes adherens junctions microvilli + + Na - K ATPase regulate the entry and exit of Na and K ATP synthase protein membrane that facilitates ATP synthesis Carbohydrate chains act as receptors A. Integral proteins(amphipathic) traverse the membrane B. Peripheral proteinsis polar (hydrophobic) C. Transmembrane proteins IV. Fluid mosaic model Lipid bilayer A. Lateral diffusion very fast (same layer) B. Transverse movement (flip flop) very slow (different layer)

V. Membrane transport Hydrostatic pressure is directly proportional to diffusion The higher the temperature, the greater the diffusion ( directly proportional) Solute o The stuff that passes through the lipid bilayer o Simple diffusion of solutes is dependent on: Concentration gradient Electrical potential Permeability coefficient Hydrostatic pressure Temperature Electrochemical gradient movement from greater diffusion to lower diffusion Movement Across Membranes Cross-membrane movement of Small Molecules Diffusion Active Transport

Cross-membrane movement of Large Molecules Endocytosis Exocytsosis

Signal Transmission Cell Surface Receptors Movement to Intracellular Receptors

Intercellular Contact and Communication Passive diffusion facilitated diffusion active transport Factors that affects diffusion concentration gradient electrical potential permeability coeficient pressure gradient temperature Endocytosis Pinocytosis cell drinking ingestion of small particles such as macromolecules types fluid-phase absorptive receptor mediated

Figure. Membrane Proteins

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Introduction to Biochemistry

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Phagocytosis cell eating ingestion of large particles such as bacteria, and other cells mostly done by Macrophages Exocytosis release of molecules from within the cell to the ECF Purposes replenishing of membrane proteins provide parts of the ECF cell signaling (hormones) A. Across a gradient 1) Passive transport occurs spontaneously across a membrane. Molecules will move from where the substance is more concentrated to where it is less concentrated. Simple diffusion no energy required; it enters the integral protein Simple diffusion (via channel) voltage/protein-gated channel o Voltage gated - activated by a change in the electrical potential of the cellular membrane in its vicinity. o Protein gated this type of diffusion causes a conformational change in the structure of the channel protein. This diffusion is limited to the speed at w/c the channel protein can change. Facilitated diffusion saturation, no. of carriers, transport; depends on the speed of its carriers. Only difference with simple diffusion is that this type needs a carrier. 2) Active transport Requires movement against electrochemical gradient Requires energy Facilitated by a membrane protein

Figure. Passive and Active transport ACTIVE TRANSPORT a substance is moved against a concentration gradient (from low concentration to high concentration) REQUIRES CELLULAR ENERGY intrinsic directionally movement of molecules is unidirectional

the

I. primary active transport: also called DIRECT ACTIVE TRANSPORT uses chemical energy such as ATP transport is linked to the hydrolysis of ATP or other high-energy molecule; for example, the + + Na /K ion pump Movement of molecules against a gradient directly linked to hydrolysis of high-energy yielding molecule (e.g. ATP) Other examples: redox energy (mitochondrial membrane against their conc. Gradient) and photon energy (light) photosynthesis that uses the energy of photons to create a proton gradient across the thylakoid membrane and also to create reduction power in the form of NADPH

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Introduction to Biochemistry

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II. secondary active transport: also called COUPLED TRANSPORT or COTRANSPORT use of an electrochemical gradient + driven by H gradient occurs when endergonic (uphill) transport of one solute is coupled to the exergonic (downhill) flow of a different solute that was originally pumped uphill by primary active transport TYPES OF SECONDARY ACTIVE TRANSPORT :

One glucose (or galactose) molecule into the cell for every two sodium ions it import into the cells

B.

COTRANSPORT SYSTEM : carries two solutes in a membrane simultaneously

Transporters Carriers, permeases Active and passive transport Slower compared to ion channels Transport/ carrier kinetics As long as the solution concentration is low the transport will be quick Transport slows down at its saturation point Specificity of transporters o Stereospecificity Ligand-gated o Charges Voltage-gated

I. ANTIPORT When two substrates move in opposite directions Example : sodium-calcium exchanger or antiporter, which allows 3 sodium ions into the cell to transport one calcium out II. SYMPORT Two substrates are moved in simultaneously in the same direction Uses downhill movement of one solutes species from high to low concentration to move another molecule uphill from low to high concentration (against electrochemical gradient) Example: o Glucose symporter SGLT1 (cotransports

Figure. Transport kinetics

Major Types of ATP-Driven Active Transporters

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Introduction to Biochemistry

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IX. Type P-type (Phosphrylation) F-type (energy coupling factors) V-type (vacuolar) Example CA ATPase; Na -K ATPase
2+ + +

Mt ATP synthase of oxidative phosphorylation ATPase that pumpts protons into lysosomes and synaptic vesicles CFTR protein; MDR-1 protein X.

Extra-large transport Endocytosis Exocytosis Gap junctions direct transfer of small molecules between cells a. connexin channel Membrane protein diseases

ABC transporter (ATPbinding cassette transporter

C.

Ion Channels Form pores in membranes Passive only Faster compared to ion channels 1) Ionopores Too much of this may cause cell lysis Channels specific for ion movement 2) Aquaporins Channels for water movement Na+/K+ ATPase most important pump in the cell moves 3 Na extracellularly and 2 K intracellurlary provides Na gradient for secondary active transport

VI.

VII. Nerve impulse transport

VIII.

Glucose transport

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Introduction to Biochemistry

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