Highlights of Chapters 1&2

• Key discoveries and Theories

• Three Kingdoms

• Cell and Genomes

• Cell Chemistry

Fundamental questions• What is the origin of life• How does life propagate• How can a single cell form a complex organism

1859 Charles Darwin, Alfred WallaceEvolution – origin of species - natural selection fittest

selected by forces of their environmentbiological adaptation

Genes of different species are closely relatedFor instance some human genes will function in yeast and fly

Historical perspective of cell biology

1950-1960 – Golden age for cell/molecular biology

Fundamental breakthroughs – basis for todays molecular understanding of biological systems

- Structure of DNA (stores genetic information, heredity)

- Central dogma (DNA RNA Protein)

- Genetic code (universal)

- Gene regulation (when, what and how much)

1980-Present – Information age of molecular biology

Water – most abundant – 75-80% by wt inorganic ions, small organic molecules such as sugars, vitamins and fatty acids can be made or imported

Macromolecules – protein, DNA, RNA, polysaccharidesmust synthesize these

Proteins and DNA are polymers of monomeric units amino acids for proteins (20) nucleic acids for DNA (4) proteins are the workhorses (proteins are versatile) (enzymatic activity, structural proteins, transport) DNA is the master molecule

MOLECULES OF LIFE

Genetic analysis(Inheritance of characteristics)

• 1865 Gregor Mendel – Pea plant

Important characteristics of his expts– Pollination control easy– Pure strains– Defined characteristics– Large sample size

• Dominant/recessive• 2 hereditary units (genes)• Independent assortment (linked traits)• One gene copy Allele

YY yy

Yy F1

YyYy

YY

yy

F2

YyYy

X

X

1865 Breeding Experimentswith Yellow & Green Pea seeds

1953 – Modern Era of Molecular Biology

- Watson/Crick, Structure of DNA

double helix

- Chargoffs rules, G=C; T=A,

rules underlying the base pairing

- Wilkin/Franklin – X-ray diffraction pattern

helical nature, diameter, distance bet adjacent bp

RNA, genetic code

1959 – Crystal structure of protein

Structure function relationships

Cell structure – Electron microscope, cell culture

1961 - Jacob and Monod – Regulation of gene

1950 - 60 - establishment of cell culture Protein sequencing

1970 identification of specific restriction enzymes dawn of cut and paste molecular genetics advent of rapid DNA sequencing oligonucleotide (DNA) synthesis

1980 Polymerase chain reaction

1990 Genome sequencing Functional genomics Systems analysis Proteomics

Three animal Kingdoms

Eukarya

Bacteria

Archaea

Based on DNA sequence similarity Archaea are more related to humans than bacteria.

Common single cell progenitor

Prokaryotes

•Prokaryotes

DNA is not sequestered

Simple internal organization

Eukaryotes

•Eukaryotes

Have a nucleus – compartment for DNA

organelles

Cells are small

Proteins are even smallerCell volume= 3.4 X 10-9 ml

Weighs 3.5 X 10-9 grams

20% protein 7 X 10-10 grams

Average protein size 52,700 grams/mol

7.9 X 109 proteins/cell

10,000 different proteins in cell

Suggests that there are over a million copies of each protein.However, levels of certain proteins are tightly controlled.Insulin receptor 20,000 copies per cell actin 5 X 108 copies

Many proteins within the cell are enzymes

Problem: How do cells keep inside water in and keep outside water out?

All cells are surrounded in a lipid membrane

What other function can membranes serve?

Compartmentalize intracellular chemical reactions

OrganellesMitrocondria-power plants

Endoplasmic reticulum-place to make membrane proteins and secreted proteins and lipids

Golgi vessicles-further refine membrane proteinsand direct their transport to specific surfaces of the cell

Peroxisomes-remove fatty acids, hydrogen peroxide and amino acids

Lysosomes-degrade old proteins and foreign materials

The Superstructure of the Cell

Blue: DNA

Red: actin cytoskeleton

Green: tubulin cytoskeleton

DNA

4 nucleotides based-paired G=C, A=T. Watson and Crick solved structure.

DNA strand coiled around a common axis forming a double helix

Flow of genetic information

Advent of genetic organization

Chromosomes resides in the nucleus means by which genetic information is transferred number and size are constant in an organism each chromosome – single DNA molecule (plus proteins) can be considered a string of genes total DNA – genome visible during cell division

Somatic cells – diploid (2n), homologous pairs (mitosis) Germ cells – haploid (n) only one of each pair (meiosis)

fruit fly (Drosophila) – 4; corn – 10; peas – 7; humans – 23

Chromosome

One human cell has 2 m of DNA found in 46 chromosomespacked into a 0.006 mm3 nucleus

Chemical nature of the gene

Arranged as regular linear arrays

Gene order could change

Gene activity Biochemical activity

One gene - One protein

DNA contains all information

subject to variation/random change

faithful reproduction (like begets like)

underlies development of every new organism

S R S

LIFE CYCLE OF CELLS

• Steady state system in adult organism balanced system (no net growth)DNA Proteins (maintenance) DNA replication

Cell divisionCell differentiationCell apoptosis

Normal cell turnover RBC nerve cells reproductive tissues

The Cell cycle

Cell Cycle follows a regular timing mechanism

Eukaryotes; Prokaryotes have no G0

Cell division 10-20 hrs vs 20-30 min

M – mitosisG1 – first gapS - synthesisG2 – second gapG0 – growth arrestcheckpoints

MitosisMitosis – Partitions genome equally at cell division

Prophase, metaphase, anaphase, telophase

Cytokinesis, mitotic apparatus

Mitosis

(go to movies)

Meiosis

Cell Death/Apoptosis/Programmed cell death/Anoikis

• Balances cell growth multiplication• eliminates unnecessary cells (development, restructuring, damaged cells)• internal program (clock) • follows systematic events (DNA frag, membran blebbing, consumed by macrophages)• Now an important area of cancer research

Cells are organized into Tissues

• Extracellular matrix (ECM) network of proteins and polysaccharides

• Cell-adhesion molecules cell-cell contact cell-ECM contact

• basal lamina• endothelium

Body Patterning dictated by patterning genes

• program of genes specify the body plan• local interactions induce specific program• Conserved throughout evolution• axial symmetry• integration / coordination of multiple events during embryogenesis

1 2 3 genetic program cell contact soluble factors

gene expression adhesion signaling

Cell Differentiation – 200 different cell types in the body

Change to carry out a special function Marked by a change in morphology “form follows function” (examples are nerve cell vs muscle cell) creates diversity of cell types requiredExamples: fertilized eggs Organism

stem cell heart & vessels Power of DNA to orchestrate cellular change

Heart Development Requires Proper Vessel Growth and Differntiation of many different cell types

CHAPTER 2 – Cell Chemistry and Biosynthesis

Chemical concepts underlying cellular processesBasic principles of chemistry and physics directbiological processes.

No supernatural force is required for biological processes

BONDS and STABILIZING FORCESCHEMICAL EQUILIBRIUMENERGYCENTRAL ROLE OF ATPENZYMES

WATER – constitutes 70-80% ; small molecules ~ 7%

Rest - MACROMOLECULES

BUILDING BLOCKS

Amino acids Proteins

Nucleotides DNA and RNA

Sugars Complex Carbohydrates

Nucleusprotons

Electrons

Orbitals

CHEMICAL BONDS

Covalent (50-200) Noncovalent (1-5 kcal/mole)

- Strong - Weak

- sharing electrons - 3D structure

within atoms of an - inter and intra molecular

individual molecule- Strength – cooperation

- multiple, weak bonds

- transient, dynamic

Covalent Bonds

A. Atoms in biological systems

• Hold the atoms within a molecule

• Formed by sharing electrons in the outer atomic orbitals

• Forms the basis of chemical reactivity and basic shape

H C N P O S 1 4 3 5 2 2,6

• Each atom can make a defined # of covalent bonds• Depends on the number of electron in the outermost orbital and their size

• typically stable (making/breaking bonds requires energy)• energy required to break a single bond (50-100 kcal/mol) double bond (120-170 kcal/mol); triple (195 “) Examples: - phosphorous – biologically very important - esters of sulfuric acid – proteoglycans in ECM

B. Bonds are oriented at precise angles (shape)

104.5 (water, each single bond)

• dependent upon mutual repulsion of outer e orbitals• non-bonding electrons also contribute to properties/shape• double bond are more rigid (cannot rotate freely)

HO

H

D. Asymmetric carbon (common in biological molecules) a carbon atom bonded to four dissimilar atoms COOH COOH H - C - NH2 NH2 - C - H

CH3 CH3

mirror image

• Optical isomers (stereoisomers) designated D or L• Central C is called chiral carbon (alpha C)• All naturally occurring aa in proteins are L.• only D form of sugars (carbohydrates are found)• different biological activity, but identical chemical property

D-alanine L-alanine

NON COVALENT BONDS or INTERACTIONS

• Hydrogen bond• Ionic Interactions• van der Waals Interactions• Hydrophobic bond

Important for stabilizing 3D structuresInter- and Intra-molecularMultiple bonds give strengthTransient/dynamic

A. Hydrogen Bond (~ 5 kcal/mol)

• Underlies chemical and biological property of water• When H atom covalently bonded to another atom (donor,D) forms a weak association (the hydrogen bond) with an

acceptor (A) atom• Both D, A – electronegative and polar• Most D, A are N (3.0) or O (3.4)

N-H C-H O-H

• Forms the basis of solubility (hydrophilic – water loving)• More H bonds, more soluble• Standard length (0.26-0.31 nm) and directionality (linear/strong)• Stabilizing force is multiplicity• H bonding usually involves exclusion of a H2O molecule

polar nonpolar

B. IONIC INTERACTIONS

• When bonded atoms have very different electronegativilty• e- found among more electronegative atom (Na+Cl-)• no fixed orientation/angel

+vely charged ion (Cation) _vely charged (Anion)

Na+, K+, Ca2+, Mg2+, Cl-

• typically exist complexed to H2O (using the water dipole)• important biological roles (nerve impulses, muscle contraction)• very soluble and energy is released as they bind water• energy of hydration

C. Van der Waals Interactions (~ 1kcal/mol)

• non-specific attractive force is created as two atoms approach each other closely • transient / momentary fluctuations in the distribution of e generating a transient electric dipole• seen in all types of molecules (polar and non-polar**)• H bonds, ionic interactions can override VDW• Van der Waal radii – balance attraction repulsion• antigen:antibody / enzyme:substrate facilitated by their complementary shape

D. HYDROPHOBIC BONDING (force that causes hydrophobic

molecules to aggregate rather than dissolve)

• non-polar molecules (for example hydrocarbons)• no ions, no dipole moment, no hydration• Force that causes non-polar molecules to aggregate• Basic force for BIOMEMBRANE structure A phospholipid bilayer typically separates two aqueous compartments (plasma membrane and organelle memb) • Phospholipids are amphipathic (tolerant of both) molecules

Fatty acyl chains – glycerol – phosphate – alcohol

Hydrophobic Hydrophilic

Spontaneously organize into structures (micelle, liposomes, bilayer)

Impermeable to salt, sugar and small molecules

VdW interactions stabilize the close packing

This structure is very fluid

Proteins – span the phospholipid bilayer

Orient their hydrophilic ends to

The aqueous environment

CHEMICAL REACTIONS

• covalent bonds are broken and re-formed• several hundred different rxns may occur simultaneously in a given cell• what rxns can proceed (rate/extent) depend on multiple factors

1. concentration of reactants (initial determinant)2. catalyst3. pH, pressure, temperature

Chemical Equilibrium: is reached when the rates of forward and reverse reactions are equal.

Equilibrium constant is the ratio of products to reactants

A catalyst can increase the rate of reaction.

A + B X + Y

Keq = [X] [Y] [A] [B]

pH: Concentration of positively charged (H+) ions

• dissociation products of H2O (H+, OH-) are constantly liberated

• when H+ is produced, it combines with a H2O molecule (hydronium ion - H3O+)

• dissociation of water is a reversible rxn

H2O H+ + OH-

@ 25o C [H] [OH] = 10-14 M2 In pure water [H] = [OH] = 10-7M

•pH = -log [H] = log 1

[H+]

•In pure water @ 25o C, [H+] = 10-7 M

•pH = -log 10-7 = 7 (Neutral)

higher value than 7 is basic;

lower than 7 is acidic

•pH – is an important property of a biological fluid

•Different cellular organelles have selective pH

•Maintenance of precise pH is imperative for cellular function

•Change in pH – a way of controlling cell activity

ACIDS and BASES

- Acid, any molecule that releases H+

- Base, any molecule that combines with H+

- organic molecules are acidic (COOH) produce COO-

O OX-COOH X-C X-C + H+

H O

X-NH2 + H+ X-NH+

3

- Whenever add acid, increase in H+

add base, increase in OH- or decrease in H+

- All solutions contain some H and OH-Biological molecules can have both acidic and basic groups-pH determines the degree to which H/OH groups are released

= =

-

COOH

H - C - NH2 @ pH 7.0

R

Amino acid

COO-

H - C – NH3+

RZwitter Ions (neutral)Doubly ionized form

pH COOH

H - C – NH3+

R

pHCOO-

H - C – NH2

R

Molecules have multiple acidic/basic groups

HA [H+] + [A-]

Ka = [H+] [A-]

[HA]

log Ka = log [H+] + log [A-]

[HA]

pH = pKa + log [A-] [HA]

pKa is the pH at which 50% of molecules are dissociated, the other 50% being neutral (Henderson Hasselbalch Equation)

• pH must be maintained near 7.2 in the cell cytoplasm• buffers are weak acids or bases (soak up [H+] and [OH-] ions• ability of a buffer to minimize the change in pH (buffering capacity)

• pKa shows the buffering capacity

Example is phosphoric acid (3 groups capable of dissociating) O

H3PO4 HO – P – OH H2PO4- + H+ pKa = 2.1

OH H PO42- + H+

pKa = 7.2

PO43-

+ H+ pKa = 12.7

Physiologically important buffer (cytosol pH 7.2, blood 7.4)=

ENERGY – defined as the ability to do work

• Kinetic (the energy of movement) - Heat/thermal; Radiant- photons**; Electric - electrons• Potential (stored energy) - Chemical bonds; Concentration gradient; Electric

potential - Important in biological systems - Glucose is the central molecule

• The law of thermodynamics: - Energy is neither created nor destroyed - converted from one form to another - Unit: Calorie (cal) = 4.18 Joules

1000 cals = 1kcal

O- – P- O – P- O – P – O – H2C Base

Sugar

= = =O O O

O- O- O-

ATP – Adenosine triphosphate (Ap~p~p)(the cellular currency for energy)

Phosphoanhydride bonds(High energy bonds) = -7.3 kcal/mole, moderate Package (easy to make, can drive many rxns) • Captures and transfers energy• used to transfer P to one of the reactants (high energy intermediate)• difference in energy released from ATP vs AMP

WHAT CAN THE ATP BE USED FOR:

• macromolecular synthesis

• cell movement (muscle contraction)

• transport molecule in/out cell

• generate concentration gradients

• generate electric potential (nerve impulse)

ENZYMES:

• straining of covalent bonds• excitation of e-

• overcome mutual repulsion of e- cloud• In biological systems kinetic energy of colliiding molecules is insufficient

• act primarily by reducing the activation energy• facilitate movement of H atoms / e- / protons• strain bonds and stabilize transition state• formation of covalent bonds

• Proteins, highly specific substrates• catalysts do not change themselves