Glycoproteins

Question 1

“Sugar Code”

Answer 1

1. Sugar (NOT GAG’s) can impart individuality to a protein because of the diversity of sugars. This diversity can include the following:
   
   1. Different number of carbons in sugar molecule (usually 3 to 9)
   2. Different types of sugars bearing specific number of carbons (heterogeneous)
   3. Different arrangement of sugar molecules
   4. Branching of sugar molecules
   5. Different types of glycosidic linkages
   6. Different types of anomers
   7. Different modifications of sugar molecules (addition of sulfate groups, phosphate groups, etc.)
   8. Different points of attachment to proteins
   9. Etc.
2. All proteins are assumed to be glycoproteins since they have at least some sugas on them.
3. There are over 1 quadrillion ways to produce possible linear and branched 20-mers of hexoses
4. The study of sugars in biology is known as glycobiology.
5. The complement (entirety) of sugars found in an organism is known as its glycome (like in genome)
6. A major class of biomolecules-bearing-sugars is known as Glycoproteins. They are NOT the same as glycosaminoglycans.

Question 2

Glycoprotein sugars

Answer 2

Again, glycoproteins are NOT GAGs

1. Glycoprotein sugars do NOT have serial repeats
2. Glycoprotein sugars are usually branched
3. Glycoprotein sugar number is less than 15 residues
4. Glycoprotein sugars are found in highly variable amounts (IgG is 4% carbohydrate by weight, while human gastric glycoprotein is 82% carbohydrate by weight); huge variation
5. Glycoprotein sugars are covalently linked to protein (similar but not the same as GAG’s need the help of linkage region for proper binding)
6. The major sugars found in human glycoproteins include the following:
   
   1. Xylose (pentose)
   2. Galactose (hexose)
   3. Glucose (hexose)
   4. Mannose (hexose)
   5. Fucose (deoxyhexose)
   6. N-Acetylgalactosamine (aminohexose)
   7. N-Acetylglucosamine (aminohexose)
   8. N-Acetylneuraminic acid (9 carbon atoms)
7. Carbohydrates can attach to the protein in an N-glycosidic link to asparagine, or in an O-glycosidic link to a serine or a threonine (both with thiol group)
   
   1. One exception is the hydroxylysine, which can be glycosylated during collagen synthesis process

Question 3

N-linked oligosacchardies

Answer 3

1. There are 2 basic classes of N-linked oligosaccharides
   
   1. Complex oligosaccharides, bearing a variety of different sugars
   2. High-mannose oligosaccharides, consisting chiefly of mannose
   3. Both the complex and the high-mannose oligosaccharides contain the same core pentasaccharide

Question 4

Functions of glycoproteins

Answer 4

1. The broad functions of glycoproteins include the following:
   
   1. Playing an essential role in the folding of the associated protein into its correct tertiary structure by possibly increasing the hydrophilicity of the proteins, to which they are covalently attached
   2. Modulating certain physical and chemical properties such as conformation, charge, binding sites, and recognition.
2. The “Glyco-“ bestows individualities to ligan and receptors found on cells
3. A subset of cell surface recognition–Antigen (ANTIbody+GENerator)–are seen as foreign, thus leading to antibody production;
   
   1. Antigenicity – the capacity to stimulate the production of antibodies or the capacity to react with an antibody.
4. Extracellular matrix, e.g. Collagen, GAGs, etc.
5. Mucins, i.e. involved in lubricating covering surfaces for physical protection

Question 5

Representative Specific Glycoprotein functions (RSGF)

Answer 5

1. RSGF associated with Lubrication/protection, Transport, Antibiotics, Structure, Recognition, Blood antigens, Hormones, Enzymes, Fertilization, and Toxins
2. Lubrication/Protection
   
   1. Mucins (“mucous”)
   2. O-linked glycoproteins found on the surface of epithelial cells of gastrointestinal, reproductive, and respiratory tracts
   3. Their high viscosity is caused by their extended structure
   4. They form protective barriers on epithelial surfaces
   5. They might possibly contain or mask certain antigens on cell surface e.g. cancer cells
   6. Lubricin found in synovial fluid (egg white)

Question 6

RSGF-Transport

Answer 6

1. Transferrin: A plasma glycoprotein that transports iron
2. Ceruloplasmin: a glycoprotein that carries most of the copper in the blood
   
   1. Lysyl oxydase require copper
   2. Wilson’s disease = genetic disease of copper metabolism
3. Serum alumin is a notable exception. It is the most abundant blood plasma protein, possessing many function including carrying a wide range of different molecules
4. Serum albumin is NOT a glycoprotein, although it is the most abundant blood proteins

Question 7

RSGF-Antibiotics

Answer 7

1. Some glycoproteins serve as non-ribosomal peptides possessing antibiotic activity; they come in many classes where some are glycoproteins in order to bind to an appropriate target

Question 8

RSGF-Structure

Answer 8

1. Collagen: a super family of structural proteins which constitutes the most abundant protein in the human body
   
   1. Collagen is part of the ECM
   2. Glycoproteins are associated with the proper folding of the collagen to make sure that the “pro”’s are not cut inside the cell.

Question 9

RSGF-Recognition

Answer 9

1. Hemagluttinin: is an antigenic glycoprotein found on the surface of the influenza virus
   
   1. Hemagluttinin allows the virus to identify target cells and to enter the cells
2. Neuraminidase is found on the surface of influenza virus.
   
   1. The neuraminidase cleaves sialic acid residues on the cell surface of proteins, allowing the virus to be released from the cell and to ultimately spread.
3. The above names (hemagluttinin/neuraminidase) are employed in the identification of specific viral strains ( e.g., H1N1, etc.)
   
   1. H_N_ denotes different kinds of hemagluttinin and neuraminidase, each for entering and leaving cells respectively; BOTH are glycoproteins
4. CD4/CD8: these transmembrane glycoproteins (Cluster of Differentiation 4,8) are essential in the activation of T-cells
   
   1. HIV binds to CD4 molecules (acting as receptor for HIV), enters these cells, and can reduce the number of CD4 T-cells
5. The above virus-cell interaction is made possible by gp (glycoprotein) 120, a protein on the surface of the HIV envelope
6. Glycophorin-C: helps erythrocytes maintain their shape
   
   1. Serves as the receptor for the erythrocyte binding antigen of Plasmodium falciparum, the infectious agent causing malaria
7. ICAM-1 (intercellular adhesion molecule-1): a glycoprotein on the surface of immune cells and endothelial cells. It can be utilized by the rhinovirus to gain access to cells; induce cold. Since they are very heterogeneous, we can’t get immune to it.
8. We have these receptors that allow for these viruses and etc. to bind to not because that was intended but because they evolve really quickly and done so to recognize the receptors of the human cells; humans ‘evolve’ relatively slowly

Question 10

RSGF-Blood antigens

Answer 10

1. The antigens (recognized by antibodies) in the ABO system are O-linked glycoproteins. The terminal sugar in these glycoproteins determines whether the antigen is AB
   
   1. The MN blood antigens are glycoproteinse each recognized by anti-M or anti-N antibodies.

Question 11

RSGF-Hormones

Answer 11

1. Thyroglobulin: serves as the precursor for thyroid hormones, T₃ and T₄
   
   1. Individuals with thyroiditis sometimes display antibodies directed against thyroglobulin
2. Erythropoietin is a glycoprotein hormone
   
   1. Erythropoietin induces the bone marrow to increase the production of erythrocytes
   2. Erythropoietin release can be stimulated by bleeding, high altitudes, etc.
3. (Human) Chorionic gonadotropin: is a glycoprotein produced by the developing embryo/placenta
   
   1. Measurement of HCG levels can serve as a test to determine pregnancy
   2. Measurement of HCG can also possibly indicate presence of certain cancers

Question 12

RSGF-Enzymes

Answer 12

1. A wide variety of different enzymes are glycosylated

Question 13

RSGF-Fertilization

Answer 13

1. Glycoproteins on the surface of sperm are essential for:
   
   1. Recognizing & targeting for the egg
   2. Evading the immune response inside of both the testes and the female

Question 14

RSGF-Toxins

Answer 14

1. Ricin: is a ribosome-inactivating glycoprotein derived from the castor oil plant
   
   1. Ricin is extremely toxic
   2. Ricin might possess potential for treating lymphoma

Question 15

Lectins

Answer 15

1. Lectins are proteins that reversibly bind specific sugars with high specificity. They have no enzymatic activity
2. Certain lectins (calnexin, calreticulin) play an essential role in glycoprotein folding
3. Lectins perform recognition on the cellular and molecular level and play numerous roles in biological recognition phenomena involving cells, carbohydrates, and proteins. Lectins also mediate attachment and binding of bacteria and viruses, as well as, mediate the first-line defense against invading microorganisms

Question 16

Synthesis of Glycoproteins/ Protein Trafficking

Answer 16

1. The glycosyltransferases involved in the elongation of the glycoproteins are attached to the membrane of the Golgi
2. They do NOT utilize a template for the synthesis of the oligosaccharides
3. The enzymes utilize nucleotide sugars i.e. UDP-derivatives (activated form of the monosaccharides) as building blocks for elongation
4. Cytoplasmic proteins are synthesized on free ribosomes, which are bound to the cytosolic side of the membrane of rER
5. Proteins that will be incorporated into the cell membrane, lysosomes, or export from cell are synthesized on ribosomes attached to the rER (rough endoplasmic reticulum) and secreted into vesicles from the Golgi
6. These proteins contain a hydrophobic signal sequence on the N-terminal, which acts to target the protein.

Question 17

Protein Trafficking Steps

Answer 17

1. The proteins are extruded into the lumen of the RER
2. The proteins are encapsulated in secretory vesicles
3. The proteins are transported to the Golgi
4. Glycolsylation occurs in the Golgi
5. The proteins are sorted out in the Golgi
6. Proteins destined for secretion or for incorporation into the lysosome are packaged in vesicles that fuse with the cell membrane or lysosomal membrane, and subsequently release the glycoprotein contents
7. Proteins destined for the cell membrane are integrated into the membrane of the secretory vesicles which buds from the Golgi and fuse with the cell membrane

Question 18

Synthesis of N-linked Glycosides

Answer 18

1. The basic process of N-linked glycoprotein synthesis is a little bit different from the O-linked process.
2. Dolicholis a membrane lipid of the endoplasmic reticulum about 100 carbons in length
3. An oligosaccharide of 14 subunits is transferred from dolichol-P-P-oligosaccharide to specific asparagine residues on the protein in a cotranslationalmanner in the endoplasmic reticulum
4. The protein bound oligosaccharide is partially processed by the removal of specific mannose and glucose residues
5. The oligosaccharide chains undergo further trimming and/or addition of monosaccharides inside the Golgi

• Sugars on dolichol get transferred to Asparagine residue of growing peptide
• Trimming are done to finally yield the final product
• Here, the process is a bulk addition instead of adding one subunit at a time
• Steps listed on the figure:
  
  1. Protein synthesis begins and the polypeptide chain is extruded into the rER.
  2. A branched oligosaccharide is synthesized on dolichol pyrophosphate.
  3. The oligosaccharide is transferred from dolichol to amide N of an asparagine residue of growing polypeptide chain.
  4. Trimming of the carbohydrate chain begins as the protein moved through the rER
  5. In the Golgi, further trimming and/or addition of monosaccharides occurs.

Question 19

I-cell disease

Answer 19

1. I-cell disease (inclusion body disease) is caused by a deficiency of N-acetyl glucosamine phosphotransferase, an enzyme normally found in the cis-Golgi. (e.g. Heinz body is also inclusion body)
2. In the absence of the enzyme, the mannose residues of nascent lysosomal enzymes are not phosphorylated (found in N-linked proteins).
3. The non-phosphorylated enzymes (lacking mannose 6-phosphate) are not targeted to lysosomes.
4. These enzymes are secreted from the cell.
5. Substrates accumulate in the lysosome creating the accumulation of inclusions.
6. The disease is considered fatal at a young age.
7. There are also genetic diseases caused by the deficiency of lysosomal exoenzymes, which function in the specific removal of one sugar associated with the glycoprotein.
8. If one enzyme is defective, degradation by the other exoenzymes cannot progress.

• Mannose residue is phosphorylated; then it will bind to mannose 6-P receptor; after which it will be able to find its way into transport vesicle which ultimately direct to lysosome. This is what SHOULD happen
• In I-cell disease, the phosphorylation never takes place.
• One defect leads to absence of ALL the lysosomal enzymes; virtual absence of all lysosomal enzymes
• Mutated phosphate transferase leads to breakdown of generation of lysosomal enzymes
• This is different from most other disorders since one defect negatively affect whole bunch of other proteins, not just one directly downstream.
• Lysosomal exoenzymes are responsible for removing terminal sugars, alluding to mucopolysaccharidoses, a disorder with enzymes breaking down certain GAG’s

Question 20

Glycoprotein summary

Answer 20