Proteins and Nucleic Acids

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Proteins and Nucleic Acids

• Also featuring amino acids and polypeptides..

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Nucleic Acids

• DNA and RNA are informational molecules in
  eukaryotic cells.
• They are built of nucleotides (the purines
  guanine and adenine and the pyrimidines cytosine,
  thymine and uracil) with a sugar-phosphate
  backbone.
• DNA has C-G and A-T RNA has C-G and A-U.

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Nucleic Acids
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Polymerization to form the nucleic acid chain

• (As with other polymers, the linkage reaction
  involves hydrolysis. In this case a
  phosphodiester bond is formed.)

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Crosslinks in the complementary pairing between
purines and pyrimidines

• Hydrogen bonds stabilize the pairs of nucleotides
  in the DNA double helix.

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Transcription DNA -gtRNA

• 1. Binding RNA polymerase binding to a DNA
  promoter sequence triggers localized unwinding of
  the double helix.
• 2. RNA polymerase initiates synthesis of
  messenger RNA from one of the two DNA strands.
• Elongation of the RNA from complementary
  nucleotides occurs as the RNA polymerase moves
  along the DNA.
• RNA polymerase reads a termination sequence and
  causes the completed mRNA strand to dissociate
  from its DNA template.

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DNA transcription
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Polytene chromosomes reveal DNA regions that are
undergoing transcription in a dipteran insect.
Highly condensed DNA, known as heterochromatin
Arrows indicate the condensed X chromosome (Barr
body) at the periphery of the nucleus of a female
mammal.
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Posttranscriptional Processing

• After a primary transcript mRNA has been
  completed,
• Introns segments of the primary transcript that
  will not contribute to the protein sequence are
  clipped out.
• The remaining exons are spliced together to form
  the mature mRNA.

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Posttranscriptional processing of the mRNA for
the beta chain of hemoglobin
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Splice variants

• In many cases, the exons can be cut and spliced
  in a number of different ways leading to splice
  variants. Splice variation is one way in which a
  single gene can give rise to multiple distinct
  proteins. Gene splicing is observed in high
  proportion of genes. In human cells, about 40-60
  of the genes are known to exhibit alternative
  splicing.

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Mechanisms that cause splice variants

• There are several types of common gene splicing
  events.
• Exon Skipping This is the most common known gene
  splicing mechanism in which exon(s) are included
  or excluded from the final gene transcript
  leading to extended or shortened mRNA variants.
  The exons are the coding regions of a gene and
  are responsible for producing proteins that are
  utilized in various cell types for a number of
  functions.
• Intron Retention An event in which all or part
  of an intron is retained in the final transcript.
  In humans 2-5 of the genes have been reported
  to retain complete introns and about 95 retain
  small parts of introns.
• Alternative 3' splice site and 5' splice site
  Alternative gene splicing includes joining of
  different 5' and 3' splice site. In this kind of
  gene splicing, two or more alternative 5' splice
  site compete for joining to two or more alternate
  3' splice site.
• The consequences of alternative splicing may be
  trivial, functional or pathological.

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(No Transcript)
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Its time to shift from nucleic acids to amino
acids, peptides and proteins
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The relationship between nucleic acids and
proteins was forged over a period of time in the
dim past, and recent appreciation that some
mitochondria, protoza and the Archaea differ from
multicellular animal cells in details of the
triplet code amino acid specification attest to
splits in the ancestry during the period of
experimentation.
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Why only 20 amino acids?

• There are at least 70 different amino acids, most
  of which are not found in proteins.
• At some very early state of evolution of life, a
  commitment to use only L amino acids was made.
  This cuts the number of possibilities by half.
• Some of the forbidden L amino acids are toxic.

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The nature of amino acids

• The building blocks of peptides and proteins can
  be divided into four categories, based on how
  they will interact within the protein
  structure/aqueous medium.
• Nonpolar (10) do not interact with water-
  located in interior of soluble proteins, or on
  exterior of intramembrane domains of membrane
  proteins
• Polar (5) hydrophilic, located in protein
  exterior for soluble proteins, or in interior of
  intramembrane domains of membrane proteins
• Basic (3) hydrophilic, form hydrogen bonds with
  water
• Acidic (2) very hydrophilic, usually located on
  the protein surface

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Amino acids
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A different text gives a different breakdown 9
nonpolar, 6 polar, etc. It is cysteine that
seems to not know where to go... .
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Amino Acid Functions

• Singaling molecules e.g. glycine, GABA (a
  glutamine derivative) and dopamine (a tyrosine
  derivative) are neurotransmitters
• Metabolizable for energy
• Sources of amine group in synthesis
• Peptide and protein subunits

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Peptides

• Small chains of amino acids (fewer than 40
  amino acids the smallest is 3 amino acids)
  serve as peptide hormones and neurotransmitters.
  Examples
• Oxytocin 9 amino acids CYIQNCPLG (C's are
  disulfide bonded). Uterine contraction, causes
  milk ejection in lactating females, responds to
  suckling reflex and estradiol, lowers steroid
  synthesis in testes
• Vasopressin antidiuretic hormone, ADH) 9 amino
  acids CYFQNCPRG (C's are disulfide bonded)
  Responds to osmoreceptor which senses
  extracellular Na, blood pressure regulation,
  increases H2O readsorption from distal tubules in
  kidney
• Melanocyte-stimulating hormones (MSH) a peptide
  13 amino acids, b polypeptide 18 amino acids,
  g polypeptide 12 amino acids. Pigmentation

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Polypeptides are longer strings of amino acids
(typically between 100 and 1000 peptide residues)

• Amino acids are joined by peptide bonds between
  the alpha amino acid group (N terminus group) of
  one amino acid and the alpha carboxyl group (C
  terminus group) of the next amino acid.
• Peptide bonds are formed in a dehydration
  reaction.
• The sequence of amino acids is listed from the
  amino to the carboxyl end

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Translation Three steps in protein synthesis

• 1. messenger RNAs
• (mRNAs) code for
• the amino acids of a
• polypeptide based
• on a triplet code.
• There are 3 stop codons and one initiation codon.
• (Note that there is not a 11 match of triplets
  and amino acids there are 64 triplets and only
  20 amino acids, so the code is redundant some
  amino acids are coded for by multiple triplets.)

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Translation, cont.

• 2. Transfer RNAs (tRNAs)
• recognize amino acids (actually, a specific
  enzyme is required to make the attachment) and
  bring them to
• ribosomes, where they line them up based on
  their complementarity to the mRNA GUG with CAC,
  etc.

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Translation, cont.

• 3. mRNA is translated into polypeptides on
  ribosomes, which are composed of ribosomal RNA
  (rRNA) and ribosomal proteins.

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Protein Structure Four levels of organization
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Chaperones Protein folding is generally not
spontaneous

• While the sequence of nucleotides in DNA is being
  translated into a sequence of amino acids to form
  a protein, the charged groups in the chain of
  amino acids are interacting, folding to allow
  to meet , or hydrophobic groups to cling
  together to avoid water. Some proteins assume
  their tertiary shape spontaneously, but in
  others, this process is assisted by chaperones,
  proteins that temporarily stabilize the
  incomplete protein by blocking associations that
  would interfere with the bending pattern
  characteristic of the functional protein.

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Diseases related to protein misfolding,
aggregation and precipitation

• Alzheimers Disease Plaques of ß amyloid result
  from aggregation and precipitation of partially
  folded 40-residue protein fragments the presence
  of a misfolded fragment initiates aggregation of
  similar fragments.
• Bovine spongioform encephalopathy (mad cow
  disease) and the related scrapie in sheep and
  Creutzfeldt-Jakob in humans are caused by prions.
  The prion protein normally plays a function in
  synapse modification in learning, but the
  abnormally folded proteins form insoluble fibrous
  aggregates.

Prion protein
Normal Diseased
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Protein misfolding occurs frequently in normal
cells

• Chaperones dont always prevent misfolding
• Misfolded proteins are labeled for destruction by
  mechanisms that are involved in turnover of
  appropriately folded proteins (i.e. proteasomes
  and lysosomes, which you will hear about in a
  subsequent lecture)

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Protein Functions

• Enzymes Catalysts
• Regulation transcription factors, protein
  hormones
• Transport hemoglobin for O2, membrane transport
  proteins
• Storage Ferritin can carry 4,500 iron molecules
• Contraction actin, myosin, tubulin
• Structural keratin, collagen
• Protective blood clotting, IGGs

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Posttranslational Processing

• In posttranslational processing, parts of the
  protein structure are removed. It may be as
  simple as removal of the signal sequence that
  directs a protein to be secreted, or as complex
  as what happens to proopiomelanocortin (POMC) to
  yield adrenocorticotrophic hormone (ACTH) (shown
  in the next slide).
• Like posttranscriptional processing,
  posttranslational processing enables one gene to
  have multiple products, violating the one gene
  one protein rule that used to be a central dogma
  of molecular biology.

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Post-translational processing of
proopiomelanocortin (POMC). POMC in mammals
consists of 3 exons, of which exons 2 and 3 are
translated. Prohormone convertases 1 and 2
(PC1/2) break the parent POMC peptide into
successively smaller peptides by cleavage at
paired dibasic amino acid residues consisting of
lysine (K) and/or arginine (R). The final
products are generated in a tissue specific
manner, for example a-MSH and ACTH are not
produced by the same cells in the pituitary. The
final products include the melanocortins (MSHs
and ACTH), ß-endorphin (ß-end) and
corticotrophin-like intermediate peptide (CLIP).
There are intermediate peptides whose biological
function remains unclear, such as ß and ?
lipotrophins (ß-LPH, ?-LPH). Millington,
Nutrition Metabolism 2007, 418.
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POMC is a single gene product that regulates many
body activities

• Depending on the type of neuron that is
  expressing it, POMC is converted to different
  endproducts that regulate a wide variety of
  activities, including feeding, stress responses,
  metabolism, pain perception, body pigmentation,
  sexual behavior, lactation, etc.

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Insulin is another example of a protein that is
created by postranslational processing
In this process, the immature protein is secreted
into the endoplasmic reticulum as it is
synthesized. Within the ER, the signal sequence
is then removed. The segments of the original
gene product that will become the alpha and beta
chains of the final hormone are folded by a
chaperone protein and inked by disulfide bonds to
form proinsulin. Removal of the C-peptide loop
connecting the two chains results in the final
hormone.
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Summary

• Nucleic acids store information (DNA) and
• transfer this information into proteins (mRNA,
  tRNA, rRNA)
• Proteins are the most diverse class of molecules
  in cells their roles are to execute the
  information carried in the DNA
• Protein function depends on protein structure.
  The 1o structure of a protein is established by
  posttranscriptional and posttranslational
  processing, and provides the basis for higher
  structural orders, which generally cannot be
  attained without the assistance of other proteins.