TOPIC 2: BIOMOLECULE 1 DNA & PROTEIN
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Transcript of TOPIC 2: BIOMOLECULE 1 DNA & PROTEIN
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TOPIC 2: BIOMOLECULE 1DNA & PROTEIN
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Genetic information
Deoxyribonucleic acid
usually
Held together by
Double helix
Single stranded
Hydrogen bonds
Denaturation/Replication
maybe
B, A, Z
Ribonucleic acid
bases
Several forms
Anti parallel strands
Primary
Structural hierarchy
sequence
Secondary Tertiary
3D confirmation Supercoiling
Linked nucleotides
Transcribed to
Made ofMade of
e.g
sugar bases Phosphate ester link
AdenosineCytosineGuanineThymineUracil
has
Can become
Stabilized by
Exist in
has
has
Is three main
rRNA tRNA mRNA
Participate in
translation
Contains code for
Protein
Is synthesis of
e.g
Histones
Condense DNA in
chromatin
e.g
e.gisis
e.g
e.g
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DNA overview
• Hallmark of life – the ability to produce• The unique information for each individual
must be preserved and passed to progeny
• All life on earth uses nucleic acids for storage genetic information
• Except for viruses; all life use deoxyribonucleic acid (DNA) to store information
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Central dogma of Molecular Biology•Sequential genetic information transferred from DNA residue to synthesis protein
•DNA play essential role in heredity by serving as template for its replication.
•DNA cannot flow directly to synthesis a protein
•Genetic information from DNA is transferred to RNA through transcription
•The sequence of RNA is translated into a protein sequence
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Structure and components of the nucleotides
• Nucleic acid consist of nucleotide monomer
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OO=P-O O
Phosphate Group
NNitrogenous base (A, G, C, or T)
CH2
O
C1C4
C3 C2
5
Sugar(deoxyribose)
Nucleic acids consist of repeating nucleotide that have phosphate ester, a pentose sugar, and a heterocyclic base.
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Nucleoside
• Base bound to pentose sugar
• Pentose sugar attached to ribose – ribonucleoside
• Pentose sugar attached to deoxyribose- deoxyribonucleosides
O
OH
N
N
NH2
O
CH2OP
O
O-
O-
deoxyctyidine monophosphate (dCMP)
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Types of nucleic acid
• DNA – Deoxyribonucleic acid• RNA – ribonucleic acid
O OHCH2
OHOH
HO HO O OHCH2
OH
ribose deoxyribose
(no O)
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Structure and component of the nucleotides
• Nucleotide also called as nucleic acid base• Base- refer to the nitrogen aromatic
compound• Nucleic acid base type: pyrimidine and purine• Pyrimidine – single ring• Purine – double ring
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Pyrimidine and Purine
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Nitrogen-Containing Bases
N
N
N
N
H
NH2
N
N
O
CH3
O
H
H
N
N
N
N
O
H
NH2
H
N
N
NH2
CH3
O
H
N
N
O
CH3
O
H
H
adenine (A) thymine (T)
guanine (G) cytosine (C) uracil (U)
Uracil generally only in RNAThymine generally only in DNA
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Nucleosides in DNA
Base Sugar NucleosideAdenine (A) Deoxyribose AdenosineGuanine (G) Deoxyribose GuanosineCytosine (C) Deoxyribose CytidineThymine (T) Deoxyribose Thymidine
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Nucleosides in RNA Base Sugar NucleosideAdenine (A) ribose AdenosineGuanine (G) ribose GuanosineCytosine (C) ribose CytidineUracil (U) ribose Uridine
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• Polymerization nucleotides form nucleic acids
• The phospodiester links the 5’OH of one residue and 3’OH of the next
• One end must terminate at in 5’OH, the other terminates at 3’OH.
O
N
N
NH2
O
CH2OP
O
O-
O-
OH
O
N
N
NH2
CH2OP
O
O-
OH
O
N
N
AMP
CMP
3
5
Β-glycosidic bond
3’-5’ Phosphodiester bonds
Formation of Nucleic Acid Structure
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Nucleic acid structure
Single letter represent individual base
Sequence of bases are unique and make each of us different!
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Double helix structure of DNA• Determination of double helix structure was
based on the X-ray diffraction patterns• Amount of T equal to A• Amount of G equal to C• Consist of 2 polynucleotide chain (we call it
DNA Strand) that wrapped to each other to form helix
• Chain run in antiparallel directions: 5’ to 3’ – sense strands 3’ to 5’ – antisense strands
• Sugar phosphate backbone – outer part• Bases pair is complementary:
A—T (2 H bond) G – C (3 H bond)
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Unwinding the helix
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Complementary base pairing
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DNA Sequence
Length of DNA sequence depends on organism:Bacteria e.g. Salmonella ~ 4Mb (4 million)Human ~ 3.4 Gb (billion)
• kb (= kbp) = kilo base pairs = 1,000 bp• Mb = mega base pairs = 1,000,000 bp• Gb = giga base pairs = 1,000,000,000 bp.
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DNA sequence• DNA sequence is obtained through Sequencing method• Sequence can be uploaded in NCBI database• Sequence of interest can also be found in the website
We will find time to review this webpage later…
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Denaturation of DNA
• A process by which double stranded DNA unwinds and separates into single stranded strands through the breaking of hydrogen bonding between the bases
• Can be achieved through heating the DNA in solution
• Complete denaturation- ~94°C
• Temperature needed depends on the base content of DNA; High G-C content will need a higher temperature. And why?
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Denaturation of DNA
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Renaturation of DNA
Reformation of complementary strands that were separated by heat by slow cooling process
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We will discuss on DNA compaction next week!
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PROTEIN 1:COMPOSITION AND
STRUCTURE
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Functional role of proteins in mammalian organism
• Catalysis in chemical transformations-enzymes• Transport –
Hemoglobin and myoglobin transport O2 in blood and muscle
Transferrin transport iron in blood• Metabolic control- enzymes involve in the process• Contraction – myosin and actin function in muscle
contraction• Matrix for bone and connective tissue – collagen and elastin
form the matrix of bone and ligament• Α-keratin- in hair and other epidermal tissue
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AMINO ACID COMPOSITION OF PROTEINS
• All different type of proteins are synthesized as polymers of only 20 amino acids
R Group- uniquely define each of amino acid
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R Groups of Amino Acids
Is used to classify amino acids:• Polar or non polar• Acidic or basic
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Polar uncharge
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Non polar hydrophobic
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Acidic
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Basic
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Abbreviations
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Amino acids can act as both acids and bases
CO2H – Can be deprotonated to become negative carboxylates (COO-) – cause acidity
NH2- – Can be protonated to become
positive α- ammonium groups (+NH3) – cause basic properties
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Amino acid as zwitterion
Basic groupAcidic group
H transfer
Zwitterion – a condition when amino acids are without charged groups on their side chain – no net charge; in solution- neutral
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Adding alkali to amino acid solution
+ [OH+]
Donate > [H+] to bind with +[OH+] NH3
+ become NH2 ( only –ve charge in COO- left)
Now this aa in negative charge!
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Adding acid to acid amino solution
+ [H+]
Now this aa in positive charge!
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Amino acid charge
General condition
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Amino acids are polymerized into peptides and proteins
• Isoelectric pH- the pH at which a molecule has no net charge – also called as isoelectric point (PI value)
• The PI- allow protein to be separated using electrophoresis, isoelectric focusing and ion exchange chromatography
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• TASK 1: Explain the function of plasma protein in diagnosis of animal disease
*Must include charge interaction and electrophoresis idea
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Two shape of proteins:• Fibrous protein• Globular protein
Fibrous protein Provide mechanical support Often assembled into large cables or threadse.g: α-keratin – major components of hair and nails
collagen – major components of tendons, skin, bones and teeth Involved in structure :tendon, ligaments, blood clots –
collagen and keratin Contractile protein in movement: muscle, microtubule
(cytoskleton, mitotic spindle, cillia, flagella)
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Globular protein• Usually water soluble, compact roughly spherical• Hydrophobic interior, hydrophilic surface• Globular protein include enzyme carrier and
regulatory protein• Most protein which move around (e.g albumin,
casein in milk)• Proteins with binding site:Enzymes, haemoglobin, immunoglobulin,
membrane receptor sites
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The peptide bond
• Peptide- short polymers of amino acid monomers linked by peptide bonds
• Polypeptide chain – longer peptide chain
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Hierarchy structure of protein
Primary structure (A.A sequence)
Secondary structure (α-helix and β-pleated sheet)
Tertiary structure (3-D structure formed by assembly of secondary structure)
Quaternary structure (structure formed by more than one polypeptide)
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Primary structure of proteins
Sequence of amino acid in polypeptide chainIs held together by peptide bondsTwo ends – N terminus and C terminus
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Secondary structure
• Local 3-D folding of the polypeptide chain in the protein
• Arrangement in space of the atoms in the peptide backbone
• Two type: α-helix and β-pleated sheet
α-helix β-pleated sheet
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• Forces involve:Strong – covalent bondWeak – hydrogen bond, electrostatic
interactions, hydrophobic effect
Secondary structure
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Tertiary structure
• 3D arrangement of all atoms in the proteins, including those in side chains and in prosthetic group
• Describes the folding and other contortions of a polypeptide chain that result from the molecular interactions among the R groups of the different amino acids
• The folding is sometimes held together by strong covalent bonds (cystein-cystein disulphide bridge)
• 3-D structure is determined through X-tray crystallography• Now can be predicted using bioinformatic technique
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Forces involved in tertiary structure
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Quaternary structure
• Arrangement of polypeptide chains in a multi chain protein
• The chain is called subunit• Subunit must be in non covalent association, maybe
connected by disulfide bonds• Not all protein have this structure• E.g.
chymotrypsin contains 3 polypeptide joined together by interchain disulfide bonds Hemoglobin – bohr effect
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DENATURATION AND REFOLDING• Non covalent interactions that
maintain 3-D structure of protein are weak – can be disrupted easily
• Unfolding a protein (i.e. disruption of tertiary structure)- denaturation
• Denaturation and reduction of disulfide bonds- happen when complete distruction of tertiary structure is desired
• Disruption process can be recovered - refolding
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Denaturation
Can be achieved in two ways:
1. Heat – increase temperature trigger vibration in molecules – when energy become great enough can disrupt the tertiary structure
2. At extreme high or low pH- some charges will be missing- so electrostatic interactions that normally stabilize the native and active form of protein are drastically reduced
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Denaturation3. Detergents – binding of detergents (e.g. sodium
dodecyl sulfate)- can disrupt hydrophobic interaction
charged detergent – disrupt electrostatic interactions
urea / guanidine hydrochloride – form hydrogen bonds with protein that stronger than internal hydrogen bond
B- mercaptoethanol – reduce disulfide bridge to 2 sulfuhydryl groups
#what do they use to straighten your curly hair?
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Relationship between protein structure and its function
• Protein structure determines protein function• Denaturation or inhibition may change protein
structure - will change its function• Coenzyme and co factor may enhance the
protein’s structure
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Protein folding• A process in which a
polypeptide folds into a specific, stable, functional 3-D structure
• In order to carry out their function (e.g. enzymes or antibodies), protein must take on a particular shape, also known as ‘fold’ – from 1° to 3°
• Thus protein are amazing machine! Before they do their work, they assemble themselves! This self- assembly is called ‘folding’
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The importance of correct folding
• Primary structure carry all the information needed to produce correct tertiary structure – but the process it self sometimes not straight fwd and trickier
• Protein dense environment cell – protein may fold incorrectly as they produced
• Or they may begin to associate with other protein before they complete their own folding
• In euk – proteins may need to remain unfolded long enough to be transported across the membrane
• Correctly folded – usually soluble in aqueous cell environment or attached in membrane
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The importance of correct folding
• Folding incorrectly – may interact with other proteins and form aggregates (Accumulate and clump together )
• Fail to do so – ineffective use of protein or producing toxic protein! – lead to protein folding disease
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• Occur because hydrophobic regions that should be buried in side the protein remain exposed and interact with other hydrophobic regions of other molecules Hydrophobic chain –
red color – interior Hydrophilic chain –green color –exterior
The importance of correct folding
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Protein folding chaperone
• Chaperone – special protein that help in correct and timely folding protein
• Prevents protein form associating with another protein or prevent it from associating with itself in inappropriate way
• E.g. of chaperone – hsp 70 (70 kilodalton heat shock protein)
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Chaperon helps in formation of hemoglobin
• Hb – consist of α globin and β globin chain
• Produced by α and β globin gene• There are 2 α globin gene for one β
globin gene – thus there is excess of α globin chain – aggregate among themselves damaged RBC – thallesemia – useless form of hemoglobin
• Therefore, α globin chain need to kept from aggregating – so they are enough in to complex with β globin chain
• With help from chaperone – α hemoglobin stabilizing protein – prevent α globin chain from causing damaged to RBC and help to deliver them to β globin chain
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TASK 2 – DISCUSS THE EXAMPLE OF PROTEIN FOLDING DISEASE BY STATING THE MECHANISM