Enzymes by Palmer: Thermodynamics and Laws

• Enzymes by Palmer is a key enzymology book
• Chapters 1, 3.2, and 6 in this book has some essential points that the text will cover

Thermodynamics

• System refers to the studied substance, while surroundings are everything else
• Closed systems exchange energy but not matter, open systems exchange both, and isolated systems exchange neither
• Physical and chemical energy types are relevant
• Cells are partially open systems that function under constant temperature and pressure
• Gibbs free energy relates to enthalpy and entropy
• ΔG represents the change in Gibbs free energy.
• Spontaneous reactions are linked to ΔG, activation energy, and reaction norm

Laws of Thermodynamics

• Zeroth Law: If system A is in thermal equilibrium with B, and B with C, then A is also in equilibrium with C
• First Law: The universe's total energy remains constant (conserved); ΔE = q + w (q = heat absorbed, w = work done)
• Second Law: Spontaneous processes increase the universe's entropy; ΔSuniverse = ΔSsys + ΔSsurr > 0 (for spontaneous reactions)
• Third Law: A pure crystalline substance at absolute zero has zero entropy: S(0 K) = 0

Entropy Equations

• Boltzmann Equation: S = k In W (k = 1.38 x 10^-23 J/K)
• ΔSsurr = -ΔHsys/T (at constant pressure)
• ΔS°sys = Σ nS° (products) - Σ nS° (reactants)
• ΔSuniverse = ΔSsys + ΔSsurr (positive for spontaneous processes)

Entropy Change in the Universe

• ΔS universe = ΔS system + ΔS surroundings
• ΔS surroundings = -ΔH system/T
• ΔS universe = ΔS system + (-ΔH system/T)
• Multiplying both sides by -T gives: -TΔS universe = ΔH system - TΔS system

Gibbs Free Energy and Spontaneity

• If ΔG is negative, a reaction proceeds spontaneously in the forward direction
• If ΔG is zero, the system is in equilibrium
• If ΔG is positive, the reaction tends to occur spontaneously in the reverse direction

Chemical Reactions and Catalysts

• A chemical reaction involves changing one or more substances into different substances.
• Reactions occur in cells constantly
• Catalysts lower the energy needed to start chemical reaction
• Catalysts increase the speed of the reaction
• Catalysts remains unchanged once its done reaction

Catalyst Effects

• Catalysts accelerate forward and reverse reactions equally, not altering equilibrium
• The oxidation of SO2 with Pt, Fe2O3, and V2O5 catalysts yields the same equilibrium composition
• Different catalysts can yield distinct product distributions from identical starting materials, impacting selectivity
• For example, ethanol decomposes into ethylene + diethyl ether with boron phosphate, or hydrogen + acetaldehyde with Mo2C/carbon

Cells and Enzymes

• Biological catalysts, largely proteins, are found in all living cells and speed up the rate of chemical reactions.
• Catalysts remain unchanged post- rxn

Biological Catalysts

• Catalase functions as a biological catalyst in plant and animal cells' cytoplasm, accelerating hydrogen peroxide breakdown

Chronology of Enzymology

• 1700s: Biocatalysis observations in the digestive tract
• 1800s: Starch conversion to sugar by saliva seen
• 1850: L. Pastour links sugar to alcohol (yeast) fermentation (vitalism)
• 1878: W. Kühne coins the term enzyme
• 1897: E. Buchner achieves sugar to alcohol conversion without cells using yeast extract
• 1926: James Sumner isolates and crystallized urease from jackfruit seeds, postulating all enzymes are proteins
• 1930: Sumner's conclusion gains wide acceptance after Northrop and Kunitz crystallize pepsin and trypsin
• J.B.S. Haldane writes Enzymes

Enzyme Nomenclature

• A nomenclature system is needed for clarity
• One enzyme, one name, one reaction; enzyme names should reflect substrate specificity
• Lactose + H2O turns to Glucose + Galactose (enzyme: Lactase)
• -O2C.CH=CH.CO2 + H2O turns to -O2C.CHOH.CH2CO2 (enzyme: fumarase)
• Trivial names don't indicate enzyme source, function, or catalyzed reaction (e.g., trypsin, thrombin, pepsin are preteases)

Need for Nomenclature

• Illustrates need for standardized names across languages: human has many names
• Human, Manav, Manava, Manitan, Manavadu, Mensch, Ningen, Menselijk, Chelovek, Rénlèi

Systematic Enzyme Names

• The International Union of Biochemistry and Molecular Biology (IUBMB) dictates a two-part enzyme naming system.
• First part: Substrates name
• Second part: Reaction type with suffix "ase" (ex. Lactate dehydrogenase)

EC Numbers

• The Enzyme Commission (EC) assigns numbers that classify the various classes enzymes, grouped by rxn
• Nomenclature determined in 1961 and updated in 1992
• Classifications dont consider amino acid sequence (homology), protein structure, or chemical mechanism
• EC numbers consist of 4 digits (a.b.c.d), where 'a' is the class, 'b' is subclass, 'c' is the sub-subclass, and 'd' represents the sub-sub-subclass
• Digits 'b' and 'c' describe the action while 'd' distinguishes between enzymes of similar purpose based on a substrate
• Ex: Alcohol: NAD+ oxidoreductase EC number is 1.1.1.1

Enzyme Classes

• EC 1: Oxidoreductases
• EC 2: Transferases
• EC 3: Hydrolases
• EC 4: Lyases
• EC 5: Isomerases
• EC 6: Ligases
• EC 7: Translocase

Oxidoreductases

• EC 1 enzymes catalyze hydrogen, oxygen, or electron transfer between substrates
• Also known as oxidases, dehydrogenases, or reductases that are 'redox' rxns needed for electron donor/acceptor

Transferases

• EC 2 enzymes catalyze group transfer reactions, minus what oxidoreductases do
• A-X + B → BX + A is general form

Hydrolases

• EC 3 enzymes catalyze hydrolytic reactions. Including esterases, lipases, peptidases/proteases, and nitrilases
• A-X + H2O → Х-ОН + НА is reaction

Lyases

• EC 4 enzymes are for non-hydrolytic removal of groups or adding functional grps

Isomerases

• EC 5 ezymes catalyze isomerization through racemizations and cis-trans isomerizations

Ligases

• EC 6 enzymes catalyze synthesis (C–X bonds), along w breakdown of containing substrates usually ATP

Translocases

• EC 7 enzymes are catalysts that encourage ion or molcule movement through the cell
• AX + B (side 1) ↔ A + X + B (side 2) moves molecules

Acids and Bases

• Brønsted-Lowry and Henderson-Hasselbalch models explain acid base
• Henderson-Hasselbalch equation

Concept of acidity

• Only A (alkaline ions) are present in presence of pH, and only HA in absence
• When [HA]=[A-], that is considered pKA
• Bronstead Lowry indicates H donors or acceptors (acids and bases, respectively)

Buffers

• Buffers are biological and can be Tris, phosphate, acetate, citrate, Hepes, MOPS

Good Buffer Criteria

• pKa is between 6 and 8
• Solubility in water must be high
• Excludes bio membranes
• Minimal salts
• has little Dissociation from changes in temperature and concentration
• Clear or non Interactions w/mineral cations.
• Stable chems
• No Light absorption
• Simple implementation

Preparing a Buffer

• Parts is Buffer component of desired pH, salts for ionic strength, reducing bME or DTT, and detergents.
• Optional parts are Glycerol, PEG, ATP, etc. that change cell reaction conditions

Amino Acids

• Building blocks of proteins and contain amine groups

Protein Buffering

• Proteins contains acidic and base groupings
• Peptide bonds are built from said group
• Protein concentration impacts molecule's pI, buffer characteristics,

Globular Protein Solubility

• Solubility varies with pH and is least soluble at its isoelectric point.

Temperature Effects

• High temperatures increase solubility
• Insoluble if salt is Zinc, Pb salt Effects insoluble Salts are trichloroacetic, perchloric, picric or sulphosalicylic acids
• Solubility decreases if non polars are present

Salting in and out

• Increasing hydrations of Salts increase the solubility
• Higher salt concentrations will decrease the proteins

Specificity of Enzymes

• From Palmer Chapter 4, Biochemistry by Voet & Voet is for refence

Enzyme-Substrate Complex

• Reversible steps are Steps involve Enzyme catalyzed Reaction
• First Step involves formation is subsrate Enzyme
• E-S Star denote Transitional period
• E-P is enzyme- product complex

Glycerol

• Glycerol becomes glycerol phosphate in enzyme substate complex

Enzyme-Substrate Complex pt. 2

• Active sites are Part of Combining Substrate w/Enzyme.
• Active sites are Has pockets, clefts, surface and R molecular groups
• Shape compliments substate shape and bonding style
• Attraction is noncovalent and conformation determines enzyme specificity

Geometric & Electronic Complementarity

• Illustrates binding points and active loops

Enzymes Specificity

• Substrates are binded via Weak interactions
• Induced fits during substrate binding are key, koschland

Noconvalent Forces

• instantaneous dipole in molecule leads induce dipoles in neighboring molecules.
• Induction is weak attraction force, VDW radii.

Crystal structure of plasmodium falciparum Fabl

• Enoyl-ACP Reductase inhibits the protein

Enzyme Specificity cont.

• enzyme reacts with only one substrate, highly specific
• Catalyzing with similar molecules that use the same functional group, low specificity
• Linkage between molecules with distinct bonds specific
• Stereochemistry recognizes two enantiomers most of time

Enzymes are Stereospecific

• Enzymes have high reactions to the sub strate with catalytic Stereoselectivity.
• Enzymes are Amino acids and have centres as active asymmetric site

Stereospecificity in substrate binding

• Hpro-Re is stereoselectivity that are catalytic, uses enzyme dehydrogenase equilibrium constants