2. 5.2 The Production of Protein
Until recently, proteins could only be made in cells.
3. 5.2 The Production of Protein
Until recently, proteins could only be made in cells.
Now small
polypeptide
chains can be
synthesized in
the laboratory.
Argonaut Technology
Quest 210 Protein
Synthesizer
4. 5.2 The Production of Protein
Overview of Protein Synthesis
Protein synthesis occurs continuously
throughout a cell’s life.
Protein synthesis is similar for eukaryotes
and prokaryotes.
Protein synthesis occurs on
ribosomes through a process
called translation.
5. 5.2 The Production of Protein
Overview of Protein Synthesis
Protein synthesis occurs continuously
throughout a cell’s life.
Protein synthesis is similar for eukaryotes
and prokaryotes.
Protein synthesis occurs on
ribosomes through a process
called translation.
6. 5.2 The Production of Protein
Overview of Protein Synthesis
Protein synthesis occurs continuously
throughout a cell’s life.
Protein synthesis is similar for eukaryotes
and prokaryotes.
Protein synthesis occurs on
ribosomes through a process
called translation.
7. Protein Synthesis in a Eukaryotic Cell.
In a eukaryotic cell, DNA is located within
chromosomes in the nucleus.
Transcription- RNA is transcribed from the
DNA template.
8. Protein Synthesis in a Eukaryotic Cell.
Introns are removed from the RNA and exons
are spliced together.
The exons comprise the mRNA (m for
messenger).
9. Protein Synthesis in a Eukaryotic Cell.
The mRNA transcripts carry the DNA code out
of the nucleus to the ribosomes, which translate
the code into a strand of amino acids.
10. Protein Synthesis in a Prokaryotic Cell.
There is no nucleus. There are no introns and
exons and the entire gene codes for protein.
Translation can begin before the mRNA is even
done being transcribed.
11. Transcription and Translation
Protein synthesis is a two-step process:
First Step:
Transcription
Genetic code must
be rewritten onto a
messenger
molecule.
RNA polymerase
attaches to the
promoter region of
a gene.
21. The GENETIC CODE
More than
one codon
for each a.a
Often the
third
nucleotide
can vary.
22. DNA MUTATIONS- Effect on proteins
Redundancy in the genetic code can
often make DNA point mutations (a
single nucleotide is changed) irrelevant.
Frame shift mutations- inserting or
deleting nucleotides that are not
multiples of threecause the greatest changes because
every a.a. after the mutation will be
wrong.
23. DNA MUTATIONS- Effect on proteins
Frame shift
mutations- inserting
or deleting
nucleotides that are
not multiples of
threecause the greatest
changes because
every a.a. after the
mutation will be
wrong.
24. Post-translational Modifications
Polypeptide chains fold into their 3D
conformations.
The protien may then be modified via
Glycosylation- addition of sugar groups
Phosphorylation- addition of
phosphate groups.
Cleavage- cut
25. Vocabulary
• Protein synthesis – the generation of new
proteins from amino acid subunits; in the cell, it
includes transcription and translation
• Transcription – the process of deciphering a DNA
nucleotide code and converting it into RNA
nucleotide code; the RNA carries the genetic message
to a ribosome for translation into a protein code
• Codon – a set of three nucleotides on a strand of
mRNA that codes for a particular amino acid
• Translation – the process of reading a mRNA
nucleotide code and converting it into a sequence of
amino acids
26. Vocabulary
• tRNA – a type of ribonucleic acid (RNA) that
shuttles amino acids into the ribosome for
protein synthesis
• Peptidyl transferase – an enzyme found in
the ribosome that builds polypeptide chains by
connecting amino acids into long chains through
peptide bonds
• Phosphorylation – adding phosphate groups
• Cleavage – process of splitting the polypeptide
into two or more strands
27. 5.2 Review Questions
1.
2.
3.
Distinguish between transcription and
translation.
If a structural gene’s code is “TAC GGC ATG
CCC TTA CGC ATC,” what will the mRNA
transcript be?
If the mRNA transcript from question No. 2
were translated into a peptide, what would
the amino-acid sequence of the peptide be?
28. Function of Antibody Proteins
Antibodies are proteins that recognize and bind
foreign molecules (antigens) for removal from
the body
29. Function of Antibody Proteins
Invasion by something foreign to the body
(an antigen) stimulates antibody production
by B lymphocytes (B cell).
30. Function of Antibody Proteins
Antibody proteins recognize a single shape on
an antigen called an epitope and bind there,
helping immune cells to recognize and attack
the antigen.
31. Function of Antibody Proteins
Antigens can be
•microorganisms (viruses, bacteria)
•microbial products (toxins)
•foreign proteins
•DNA and RNA molecules
•drugs
•other chemicals
32. Function of Antibody Proteins
Antibodies are also called immunoglobulins(Ig)
Most is IgG
33. Function of Antibody Proteins
Epitopes are the specific parts of antigens that
are recognized by antibodies.
•Each antibody recognizes a single epitope.
Multiple antibodies may
recognize and bind to
different epitopes on a
single antigen.
34. Function of Antibody Proteins
Epitopes are the specific parts of antigens that
are recognized by antibodies.
•Each antibody recognizes a single epitope.
•Multiple antibodies may
recognize and bind to
different epitopes on a
single antigen.
35. Function of Antibody Proteins
An HIV virus
particle (virion)
has many
potential epitopes
on its surface that
may be
recognized by
many different
antibodies.
36. Function of Antibody Proteins
Structure of IgG bound to the HIV capsid protein p24
as determined by X-ray crystallography.
(Harris et al.1998, Momany et al. 1996)
37. Antibodies are mass produced via many
methods.
polyclonal antibodiesa mixture of antibodies
for a single antigen
monoclonal antibodiesclones of a single
antibody
40. ELISA
(Enzyme-Linked Immunosorbent Assay)
-a useful form of analysis that exploits the
amazing specificity of antibodies to their
antigens.
Antibodies are
designed to bind
specific molecules
and produce a
visible color.
41. ELISA
(Enzyme-Linked Immunosorbent Assay)
-a useful form of analysis that exploits the
amazing specificity of antibodies to their
antigens.
Antibodies are
designed to bind
specific molecules
and produce a
visible color.
42. ELISA
Used for detecting all sorts of molecules and organisms
(Enzyme-Linked Immunosorbent Assay)
-HIV testing or any virus
-a useful form of -drug testing
analysis that exploits the
amazing specificity of antibodies to their
-pregnancy testing
-detection of allergens (gluten, soy, peanuts)
antigens.
-identify bacteria
-detect parasites
Antibodies are
- water contaminants
designed toGMO
bind
-detect
specific molecules
ELISA can also quantify - Tells HOW MUCH
and produce a
visible color.
43. Vocabulary
• Antigens – the foreign proteins or
molecules that are the target of binding by
antibodies
• Epitope – the specific region on a
molecule that an antibody binds to
44. Vocabulary
• ELISA – short for enzyme-linked
immunosorbent assay, a technique that
measures the amount of protein or antibody in
a solution
• Monoclonal antibody – a type of antibody
that is directed against a single epitope
• Hybridoma – a hybrid cell used to generate
monoclonal antibodies that results from the
fusion of immortal tumor cells with specific
antibody-producing white blood cells (B-cells)
45. 5.1 Review Questions
1.
2.
How many polypeptide chains are
found in an antibody, and how are they
held together in the protein?
What is the value of monoclonal
antibody technology?
46. The Importance of Proteins in
Biotech R&D
The ability to synthesize and modify peptides
or proteins is crucial to the production of
virtually every biotechnology product.
47. 5.3 Enzymes: Protein Catalysts
Enzymes and Their Substrates
Enzymes are proteins that act as catalysts.
Enzymes are involved in virtually every reaction in
a cell.
Many companies have focused on producing
enzymes for sale.
48. 5.3 Enzymes: Protein Catalysts
Enzymes and Their Substrates
The molecules upon which enzymes act are
called substrates.
49. 5.3 Enzymes: Protein Catalysts
Enzymes and Their Substrates
Enzyme active site and substrate match
exactly (the Lock and Key Model)
51. 5.3 Enzymes: Protein Catalysts
Factors That Affect Enzyme Activity
Amount of substrate in the solution
Temperature
Acidity or alkalinity
Enzymes have an optimum
temperature and pH.
52. 5.3 Enzymes: Protein Catalysts
Factors That Affect Enzyme Activity
Amount of substrate in the solution
Temperature
Acidity or alkalinity
Enzymes have an optimum
temperature and pH.
53. 5.3 Enzymes: Protein Catalysts
Factors That Affect Enzyme Activity
All proteins denature in extreme temps
and outside of their optimum pH.
54.
55. Vocabulary
• Substrate – the molecule that an enzyme acts on
• Lock and key model – a model used to
describe how enzymes function, in which the
enzyme and substrate make an exact molecular fit
at the active site, triggering catalysis
• Induced fit model – a model used to describe
how enzymes function, in which a substrate
squeezes into an active site and induces the
enzyme’s activity
56. Vocabulary
• Optimum temperature – the temperature
at which an enzyme achieves maximum activity
• Denaturation – the process in which
proteins lose their conformation or threedimensional shape
• Optimum pH – the pH at which an enzyme
achieves maximum activity
57. 5.4 Studying Proteins
A technician loads protein samples on a vertical gel.
Vertical gel boxes operate in a fashion similar to
horizontal gel boxes.
58. Vertical Gel Electrophoresis. Gel cassettes are snapped or
screwed in place (right). Running buffer is added behind the gel,
covering the wells. Buffer is poured in the front of the gel cassette
to cover the front opening. When the top is placed on the box
(left) and the power is turned on, electricity flows from the top
(negative charge) to bottom (positive charge). Negatively charged
samples move down the gel toward the positive electrode.
59. Silver stain is much more sensitive than Coomassie® Blue. When
samples have low concentrations of protein or DNA, silver-staining
is the method of choice.
60. 5.5 Applications of Protein Analysis
Protein profile of cells and tissues
A protein’s structure can help explain its
function
Study chemical processes in cells
Evolutionary and taxonomic relationships
