Introduction to Cell biology

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Introduction to Cell biology
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Hierarchical organisation of the structure of
living systems
organisms
organs
tissues
cells
Nucleus, mitochondria, Golgi apparat, etc
Ribosomes, chromosomes, cytoskeleton, membranes,
etc
polysaccharides
triacylglycerols
nucleic acids
proteins
monosaccharides
phospolipids
nucleotides
N-containing bases Ribose
Fatty acids, glycerol, cholin
aminoacids
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Cells as seen before the cell theory
Anton van Leeuwenhoek, XVII. century algae,
bacteria, sperm cells, etc.
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Robert Hooke 1665 cell unit in dead samples
of cork.
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The cell theoryCell as the central unit of
biological organization

• Cells are the basic units of life.
• All living organisms are made up of cells.
• Only living cells can produce new cells.
• Matthias Schleiden 1838
  Theodor Schwann 1835
• plants are made up of cells animals are made up
  of cells

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• Rudolf Virchow 1858
• Every animal appears
• as the sum of vital units,
• each of which bears in itself
• the complete characteristics
• of life

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Louis Pasteur
1865 Spontaneous generation of life ruled out
experimentally There is now no circumstance
known in which it can be affirmed that
microscopic beings came into the world without
germs, without parents similar to themselves."
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Tranzitions from non-living towards living I.
Prions molecules resembling ion channels,
causing serious illnesses
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Tranzitions from non-living towards living II.
Viruses
Viruses have no metabolism and can not reproduce
by themselves. They contain genetic material
(either RNA or DNA) and proteins. After infection
they use the machinery of the host cell to
produce more viruses.
Highly simplified structure of a virus
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The HIV virus
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Prokaryotic and eukaryotic cells
Diagram
EM
1 mm
1 mm
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I. (BIO)CHEMICAL FOUNDATIONS
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The most important groups of organic
molecules Proteins composed of amino
acids Lipids composed of glycerol and fatty
acids Carbohydrates mono-, oligo- and
polysaccharides Nucleic acids DNA, RNAs
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I./1. PROTEINS

• Classification of proteins
• Enzymes
• Receptors
• Transport proteins
• Storage proteins (casein in milk, ferritin
  /iron/)
• Contractile proteins
• Structural proteins
• Immune proteins
• Regulatory proteins
• Others (e.g. antifreeze proteins)

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Amino acids General chemical structure NH2-knk,
,-COOH Peptide bound NH2-knk,,-COOH
NH2-knk,,-COOH i NH2-knk,,-CONH-knk,,-COOH
H2O 20 different amino acids in unlimited amount
in any possible variations may form unlimited
number of various peptide chains
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Primary structure
Primary structure or sequence linear arrangement
of the amino acids that constitute
the polypeptide chain Sequencing to
determine the order of amino acids of a
protein. Sequence motive a specific amino acid
arrangement that appears in several different
proteins and play the same role in these
proteins. Examples DNA binding motive
signal sequence (transport of the protein
to a given organelle) sequence for
phosphorylation ligand-binding sequences
(e.g. ATP, growth hormons)
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Secondary structure
Local organisation (folding) of parts of a
polypeptide chain. Most important secondary
structure elements a-helix and b-sheet (
L. Pauling, early 1950s) In the rodlike a-helix
the polypeptide backbone is folded into a spiral
that is held in place by hydrogen bonds.
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The b sheet consists of laterally packed b
strands (extended polypeptide structures). b
sheets are stabilized by hydrogen bonds between
the strands.
The compact structure of the proteins is ensured
by turns (compact, U-shaped elements stabilized
by H-bonds) and loops (long, loose bends) between
the a-helical and b-sheet structures.
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An example Ribonuclease
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Tertiary and quaternary structure
Tertiary structure Three-dimensional
arrangement of all amino acids, which results in
mainly from hydrophobic interactions between
nonpolar amino acid side-chains. These
interactions hold helices, strands and coils
together. The highest level of organisation for
monomeric proteins. Quaternary structure
number and relative positions of subunits in
multimeric proteins. Determination of the
three-dimensional structure of proteins x-ray
crystallography
nuclear magnetic resonance (NMR)
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An example Haemoglobin
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I./2. LIPIDS AND THEIR COMPONENTS
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Triacylglycerols
Serve for storage (lipid droplets in fat cells)
and isolation.
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Membrane lipids
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Cholesterol
In addition to the phospholipids, it occurs in
biological membranes exclusively in eukaryotes.
Stabilizes the membranes.
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I./3. CARBOHYDRATES
The most abundant biomolecules on the
earth. Essential components of foodstuff
(sugar) Forms of occurence in living systems
monosaccharides (e.g. glucose) oligosaccharides
(e.g. saccharose, lactose) polysaccharides
(e.g. glycogene, starch) Occurrence in complex
macromolecules with lipids (e.g.glycolipides) w
ith proteins (glycoproteins and
proteoglycans) within nucleic acids
(constituents of RNA and DNA)
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Some monosaccharides Glycogene
polysaccharide
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I./4. NUCLEIC ACIDS
Nucleic acids are the information-storing
molecules of the cells. They are linear polymers
of nucleotides connected by phosphodiester bonds.
A nucleotide is composed of
an organic base a pentose (five-carbon sugar) a
phosphate group
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The base components of nucleic acids
N-containing (heterocyclic) ring molecules
purines ( a pair of fused ring) and pyrimidines (
a single ring).
cytosine (C), adenine (A) guanine (G) in RNA
and DNA thymine (T) in DNA
uracil (U)
in RNA
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Chemical structure of nucleic acids
DNA or RNA strand formation polymerization
(condensation) of nucleotides, by forming
phosphodiester bonds.
Nucleic acid sequence with one-letter codes e.g.
A-C-T-T-C-G-G beginning with 5end
In RNA the sugar component is ribose (one OH more)
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RNA
The RNA molecule is most often single-stranded.
Intramolecular basepairs are forming frequently
(e.g. tRNA), resulting in formation of secondary
structure elements.
Further organization of secondary structures lead
to the appearance of tertiary structure.
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• A considerable fraction of RNA occurs in great
  complexes together with proteins (e.g. ribosomes)
  
• RNA can have catalytic activity (ribozymes).
• RNA is the genetic material in several viruses
  (polio, influenza, rota, HIV, etc).

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DNA its native state is a righthanded double
helix of two antiparallel chains
The bases of the two chains ( one running 5 3,
the other one 35) are held in precise register
by H-bonds.
Base-pair complementarity A is paired with T G is
paired with C
thymine
cytosine
H-bonds
guanine
adenine
sugar-phosphate backbone
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Space-filling model of the DNA double helix
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Nobel Prize 1962
Francis Harry Compton Crick Institute of
Molecular Biology Cambridge, United Kingdom
James Dewey Watson Harvard University
Cambridge, MA, USA
for their discoveries concerning the molecular
structure of nucleic acids and its significance
for information transfer in living material
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General principles of nucleic acid polymerization

• Both DNA and RNA chains are produced in cells by
  copying a preexisting DNA strand (template)
  according to the rules of Watson-Crick DNA
  pairing /A-T, G-C, A-U/.
• Nucleic acid growth is in one direction from the
  5 (phosphate) end to the 3 (hydroxyl) end.
• Special enzymes (polymerases) are necessary to
  produce DNA or RNA.
• DNA double helix synthesis by base-pair copying
  requires the unwinding of the original duplex. A
  single stranded region (growing fork) is formed.

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I.4.1. Cellular processes involving nucleic acids
Gene expression
DNA
RNA
Protein
Trans- cription
Trans- lation
Repli-cation
Cell division
DNA
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The central dogma of genetics
DNA RNA Protein
DNA stores the information RNA is the messenger
(sometimes stores information, sometimes acts as
an enzyme) Proteins are structural units and
working molecules.
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The genetic code organisation and transformation
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The genetic code (RNA to amino acids)
The genetic code is (almost) universal the
meaning of each codon is the same in most known
organism. Unusual codon usage occurs in
mitochondria, chloroplasts and several
archaebacteria.
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Reading frames
The genetic code is commaless! Thus 5___
GCUUGUUAACGAAUUA__ mRNA
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I.4.2. Gene and genome
Gene The nucleotide sequence needed to produce a
functionally competent working molecule (RNA or
protein Genome The totality of the genes of a
given organism.
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Genome Sequence Projects
Since 1995 the following complete genom sequences
became available Prokaryotes More than 30
Bakterial species (several disease-causing ones),
some Archaebakteria Eukaryotes Saccharomyces
cerevisae (bakers yeast) Caenorhabditis elegans
(worm) Drosophila melanogaster (fruitfly) Arabidop
sis thaliana (plant) Mus musculus (mouse) Homo
sapiens
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(No Transcript)
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The human genome

• the sequence of the human genom contains 3,3
  billion bases, organised in 24 chromosomes (22,
  X,Y)
• 30 000 to 40 000 genes
• 233 genes are evidently of bacterial origin
• 98 of the sequence is nonfunctional
• the genetic identity of the human beings is 99.9
  
• Nature, 15. February 2001/Science, 16. February
  2001

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1.4.3. Gene expression
Gene expression the entire cellular process
whereby the information encoded in a particular
gene is decoded to a particular protein.
Molecular processes involved in gene expression
transcription und translation. During
transcription an RNA (messenger RNA, mRNA) is
synthesized, which contains the genetic
information of the DNA as a complementary
sequence. The procedure is catalyzed by DNA
dependent RNA polymerases. During translation
the nucleotide sequence of the mRNA is converted
to amino acid sequence of a protein. Besides the
mRNA, ribosomes and tRNAs numerous enzymes and
regulator proteins play important roles in this
procedure.
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Organization of genes in DNA in prokaryotes and
eukaryotes
Prokaryotes Protein-coding regions, organized
in operons, are closely spaced along the DNA
sequence. Example the lac operon of E. coli
(Jacob and Monod, 1960s)
Transcription control region
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Main features of gene expression in prokaryotes
and eukaryotes
Prokaryotes Example lac operon
RNA polymerase
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Eukaryotes a considerable amount of DNA is
untranslated Transcribed regions of most of the
genes is composed of several exons (translated
from mRNA) and introns (eliminated from mRNA
before translation). Example human beta globin
gene
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90
130
222
850
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Untranslated regions Exons Introns
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Eukaryotes

• Trancription occurs in the nucleus, translation
  in the cytoplasm.
• Primary RNAs undergo processing within the
  nucleus ? addition of 5cap
  ? polyadenilation ? splicing
  (removal of introns)
• mRNAs are monocistronic.
• Besides the nucleus, DNA occurs also in
  mitochondria and chloroplasts.

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1.4.3.1. Transcription
Catalyzed by DNA dependent RNA polymerase. Steps
of the procedure 1. the RNA polymerase finds an
appropriate initiation site on the duplex DNA and
binds to it
2. The enzyme temporarily separates the two DNA
strands
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3. De novo RNA synthesis begins by the binding
of the first nucleotide by base pairing
4. The second nucleotide binds by base paring.
The enzyme catalyses the linkage of the two
nucleotides (PPP remains at the 5 end, PPi is
split off from the second nucleotide).
5. The third nucleotide binds and the enzyme
links it to the existing dinucleotide. The
procedure continues until the STOP codon.
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1.4.3.2. Translation.

• Participants
• mRNA source of the genetic information
• loaded tRNA adaptormolecule, recognizing the
  codon and providing the corresponding amino acid.
• Ribosomes the machines in which the
  proteins are produced on the basis of the
  genetic information provided by mRNA.
• Numerous other proteins serving as regulators
  intiation- elongation and terminationfactors
• GTP and ATP

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Loaded tRNA
Function to furnish the appropriate amino acid
on the basis of the code on the mRNA.
Base pairing
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The ribosome
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Molecular components of ribosomes
Large subunit Mw 2 800 000
Small subunit Mw 1 400 000
50 proteins 3 rRNAs
33 proteins and 1 rRNA
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The steps in translationA. Initiation
a.) a partial initiation complex forms
Met-tRNAmet binds to the small ribosomal
subunit b.) the above complex binds to the
initiation site on mRNA AUG (codon of Met) c.)
by binding of the large subunit the initiation
complex is ready to begin the synthesis
ATP and GTP is hydrolyzed and numerous proteins
initiation factors take part in these processes.
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B. Elongation
P site outgoing site. Direction of the
ranslocation of ribosomes on mRNA 5 ? 3 GTP
is hydrolyzed, elongation factors take
part Elongation proceeds until STOP signal
reached.
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C. Termination
When the ribosome arrives to the stop codon (UAG)
the translation is completed
hydrolysis of peptidyl-tRNA on the ribosome
release of the completed polypeptide and the last
tRNA
dissociation of the ribosomal subunits
Termination factors play a role in the process.
GTP is needed.
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Free and ER-bound ribosomes
mRNA encoding a
cytosolic protein
pool of ribosomal subunits in cytosol
mRNA encoding a protein targeted to ER
ER membrane
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Peptide synthesis on ER-bound ribosomes
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Posttranslational modification of proteins

• After the completiton of translation numerous
  polypeptides and proteins undergo
  posttranslational modifications. These
  modifications can influence their structure and
  function. Most important posttranslational
  modifications
• specific proteolysis
• removal of the first Met
• glycosylation
• phosphorylation

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1.4.4. DNA replication
Semiconservative replication every double helix
contains a parent strand and a newly synthetised
one.
Parent First generation Second generation
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Synthesis of the complementary daughter DNA
strands
DNA polymerases carry out DNA synthesis on a DNA
template, exclusively in 5 to 3 direction.
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Daughter duplex
5
Parental DNA duplex
5
3
3
5
3
Lagging strand
Daughter duplex
5
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DNA polymerases are unable to initiate de novo
DNA synthesis, but can add nucleotides to the
3end of preexisting RNA or DNA strands (RNA
primer, synthetised by the enzyme primase).
Leading strand DNA synthesis is continuous
Leading strand template
5
3
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Lagging strand DNA synthesis is discontinuous
Lagging strand template
5
3
5
3
Okazaki fragment (200 nucleotides)
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DNA repair, mutations
Maintainig genetic stability requires accurate
mechanism of replication as well as repair of
lesions that occur continually in DNA. Most
spontaneous changes are immediately corrected by
the complex process of DNA repair. DNA repair,
similarly to replication, relies on base-pairing
and involves several different pathways. If this
process fails, permanent change mutation
occurs in DNA. Mutations in vital positions of
the DNA sequence destroy the organism, others
might cause advantageous modifications in the
gene products, contributing to the driving force
of the evolution.