Structure of Antibody Molecule: pgs 14-18 Flashcards
What is needed for immune serum that can kill the immunizing bacteria?
- Heat stable antibody
- Heat labile complement
Blood can be divided into two parts, the ____________________ and ___________ (or ________).
Formed elements and plasma (or serum)
____________ is the supernatant of clotted blood.
Serum
__________ is the liquid portion of blood or the supernatant of blood.
Plasma
What is the difference between plasma and serum?
Plasma still has the clotting proteins present and serum doesn’t.
T/F - Serum contains both antibodies and complement.
True
Proteins that are soluble in water are _______________.
Albumins
Proteins that are soluble in saline are _________________.
Globulins
What is a pneumonic for remembering albumins and globulins?
Albus dumbledore(Albumin) walked on water but goblins (globulins) have to sail (saline).
What is a technique whereby charged molecules in solution are separated according to their net electric charge?
Electrophoresis
Out of albumin and globulins which one moves toward the cathode/anode? Why?
- Albumin - greatest negative charge - moves toward anode
- Globulin - varies in charge, some move towards cathode - they are divided into three bands in electrophoresis (alpha, beta, gamma)
Immunoglobulins are also known as ______ globulins due to what study?
Gamma globulins - Tiselius and Kabat in 1939 - An immune serum was divided into two aliquots, one was subjected to serum protein electrophoresis (SPE) and was the control and the other was precipitated by its specific antigen, then subjected to electrophoresis. In the experimental the gamma-globulin peak was substantially reduced.
What are the sedimentation coefficients (high and low MW fraction) for gamma globulin and what study found this?
19S for high MW fraction and 7S for low
Porter and Edelman study
What does Fab stand for?
Fragment-Antigen binding
What does Fc stand for?
Fragment-crystalline (this piece would crystallize during cold storage).
How many pieces did the 7S fragment of the gamma globulin break into when it was subjected to papain?
3 pieces: 2 (45 kDa) which were the Fab portions and 1 (50 kDa) which was the Fc region
How many pieces did the 7S fragment of the gamma globulin break into when it was subjected to pepsin?
1 single 100 kDa piece, known as F(ab)’2
This piece retained its reactivity with Ag and would also precipitate Ag which Ab digested with papain would not.
The 7S Ab was subjected to electrophoresis and migrated as a single band with a molecular weight of _____ kDa.
150 kDa
How did the single 150 kDa react when it was reacted with Beta-ME and electrophoresed in 8M urea?
It was converted into two molecules:
-Heavy chain: 50 kDa
-Light chain: 25 kDa
This is why it was proposed that Ab consists of two heavy chains and two light chains
T/F - On a given antibody molecule, the chains can be both kappa and lambda.
False - they can either be kappa or lambda, but never both.
Does kappa or lambda appear more frequently in humans?
Kappa
How were scientists able to properly study and sequence antibodies?
Due to the disease multiple myeloma, which plasma cells become cancerous and secretes normal Ab at an increased level. This large enough quantities made it possible for purification. This was due to the fact that all of the myeloma protein in the patient’s serum was identical.
What type of bonds hold together the two heavy chains or heavy and light chain together?
Disulfide bonds (cysteine-cysteine bonds)
What causes the immunoglobulin portions to loop?
Intrachain disulfide bonds - approximately 60 amino acids long
The immunoglobulin loops are referred to as ____________.
Domains
How many domains does the heavy and light chain have?
- Light: 2 domains
- Heavy: 3 or 4 domains
The first domain of both the heavy and light chains was found to have a __________ structure.
variable - this contains the first 100 amino acids of the amino terminus
___________ regions or Complementarity Determining Regions (CDRs) are three areas in a variable region that are even more variable than the rest of the V region.
Hypervariable regions
What is a paratope?
Antigen binding site
What does the binding site consist of?
Variable regions from one light and one heavy chain, for a total of six CDRs per paratope
What are the non-CDR areas referred to as?
Framework regions (FR)
The isotype (class) of the antibody is determined by the __________ region.
constant
Is the VH and VL interaction stronger or weaker than the CH1 and CL interaction?
Stronger
Gamma, delta, and alpha heavy chains contain an extended peptide sequence between CH1 and CH2. This sequence is the _________ region and provides additional flexibility
Hinge
Which immunoglobulins have hinge regions and which do not:
Hinge regions: IgG, IgA, and IgD
Other regions: IgM and IgE
What is an isotype?
Ab classes and subclasses. All members have one copy (per chromosome) of each heavy chain and light chain class and subclass
What is an allotype?
Abs with minor differences in the amino acid sequence in the constant regions of the heavy or light chains within an Ab class. They are differing alleles and in humans they are denoted as Gm or Am.
What is an idiotope?
Unique sequences in the variable regions (primarily the CDRs) due to genetics.
What is an idiotype?
The collection of idiotopes (remember there are 6 CDRs per Fab). They are unique to a given individual
What are some functions of antibodies?
- Neutralize toxin/viruses
- Immobilize microbes
- Agglutinate microorganisms
- Form precipitates
- Facilitate destruction of microbes via complement
- Enhance phagocytosis
What is the reason for different isotypes?
Not every antibody functions the same. Therefore you need different antibodies to perform different functions.
