GLYCOPROTEINS

OBJECTIVES
Explain the importance of glycoproteins in health and disease.
Describe the principal sugars found in glycoproteins.
Describe the major classes of glycoproteins (N-linked, O-linked, and GPI-linked).
Describe the major features of the pathways of biosynthesis and degradation of
glycoproteins.
Explain how many microorganisms, such as influenza virus, attach to cell surfaces
via sugar chains.

BIOMEDICAL IMPORTANCE
 The glycoproteins are proteins that contain oligosaccharide chains (glycans)
covalently bound to amino acids;
 glycosylation (the enzymic attachment of sugars) is the most frequent
posttranslational modification of proteins.
 Many proteins also undergo reversible glycosylation with a single sugar (N-
acetylglucosamine) bound to a serine or threonine residue that is also a site for
reversible phosphorylation. This is an important mechanism of metabolic
regulation.
 Nonenzymic attachment of sugars to proteins can also occur, and is referred to
as glycation. This process can have serious pathologic consequences (eg, in
poorly controlled diabetes mellitus).
 Glycoproteins are one class of glycoconjugate or complex carbohydrate—
molecules containing one or more carbohydrate chains covalently linked to
protein to form among others, glycoproteins, or proteoglycans or to lipid (to
form glycolipids.
 Almost all plasma proteins, and many peptide hormones, are glycoproteins, as
are a number of blood group substances.
 Many cell membrane proteins contain substantial amounts of carbohydrate,
and many are anchored to the lipid bilayer by a glycan chain.
 Evidence is accumulating that alterations in the structures of glycoproteins and
other glycoconjugates on the surface of cancer cells are important in
metastasis.
 Glycoproteins

Proteins Oligosaccharides (<4%)

short
heterooligosaccharide
chain (branched); can be
charged; do not have
repetitive disaccharide
units
Glycosylation Characteristics of Glycoproteins
At ER Lumen At Golgi complex
Glycoproteins contains N-terminal signal sequence Glycoproteins contains O-terminal signal sequence
(N-Glycosylation ) (O-glycosylation)
Or Threonine

In this case, the proteins are sorted

Once properly made, the glycoproteins will be:
 Secreted
 Incorporated to cell membrane
 Translocated to lysosome (if impaired, the I-
cell disease  a genetically inherited lysosomal
storage disease that is caused by a defective
phosphotransferase enzyme that is located in
 the Golgi apparatus)

Glycoproteins are degraded by lysosomal enzymes (not possible if there is enzyme deficiency)
GLYCOPROTEINS OCCUR WIDELY & PERFORM NUMEROUS
FUNCTIONS
 Glycoproteins occur in most organisms, from bacteria to human beings.
o Many viruses also contain glycoproteins, some of which play key roles in
viral attachment to host cells.
 The glycoproteins have a wide range of functions; their carbohydrate content
ranges from 1 to over 85% by weight.

Some Functions Served by Glycoproteins

  The glycan structures of glycoproteins change in response to signals involved in
cell differentiation, normal physiology, and neoplastic transformation. This is the
result of different expression patterns of glycosyltransferases.

Some Functions of the Oligosaccharide Chains of Glycoproteins

OLIGOSACCHARIDE CHAINS ENCODE BIOLOGICAL INFORMATION

 The biological information in the sequence and linkages of sugars in glycans differs
from that in DNA, RNA, and proteins in one important respect;
o it is secondary rather than primary information.
o The pattern of glycosylation of a given protein depends on the pattern of
expression of the various glycosyltransferases in the cell that are involved in
glycoprotein synthesis, the affinity of the different glycosyltransferases for
their carbohydrate substrates, and the relative availability of the different
carbohydrate substrates.
o Because of this there is microheterogeneity of glycoproteins, something
that complicates their analysis. Not all of the glycan chains of a given
glycoprotein are complete; some are truncated.
o The information from the sugars is expressed via interactions between the
glycans and proteins such as lectins (see below) or other molecules. These
interactions lead to changes in cellular activity.
EIGHT SUGARS PREDOMINATE IN HUMAN GLYCOPROTEINS
 Only eight monosaccharides are commonly found in glycoproteins.
o N-acetylneuraminic acid (NeuAc) is usually found at the termini of
oligosaccharide chains, attached to subterminal galactose (Gal) or N-
acetylgalactosamine (GalNAc) residues.
o The other sugars are usually found in more internal positions.
o Sulfate is often found in glycoproteins, usually attached to galactose, N-
acetylglucosamine, or Nacetylgalactosamine.

The Principal Sugars Found in Human Glycoproteins

 As in most biosynthetic reactions, it is not the free or phosphorylated sugar that is
the substrate for glycoprotein synthesis, but the corresponding sugar nucleotide;
 some contain UDP and others guanosine diphosphate (GDP) or cytidine
monophosphate (CMP).
LECTINS CAN BE USED TO PURIFY GLYCOPROTEINS & TO
INVESTIGATE THEIR FUNCTIONS
 Lectins are carbohydrate-binding proteins that agglutinate cells or precipitate
glycoconjugates; a number of lectins are themselves glycoproteins.
o Lectins contain at least two sugar-binding sites;
o Proteins with only a single sugar-binding site will not agglutinate cells or
precipitate glycoconjugates.
o Lectins were first discovered in plants and microorganisms, but many lectins
of animal origin are now known, including the mammalian
asialoglycoprotein receptor.
 Many peptide hormones, and most plasma proteins are glycoproteins. Treatment
of the protein with neuraminidase removes the terminal N-acetylneuraminic acid
moiety, exposing the subterminal galactose residue.
o This asialoglycoprotein is cleared from the circulation very much faster than
the intact glycoprotein.
o Liver cells contain an asialoglycoprotein receptor that recognizes the
galactose moiety of many desialylated plasma proteins, leading to their
endocytosis and catabolism.
 Plant lectins were formerly called phytohemagglutinins, because of their ability to
agglutinate red blood cells by reacting with the cell surface glycoproteins.
 Undenatured lectins in undercooked legumes can lead to stripping of the
intestinal mucosa by agglutinating the mucosal cells.
 Lectins are used to purify glycoproteins, as tools for probing the glycoprotein
profiles of cell surfaces, and as reagents for generating mutant cells deficient in
certain enzymes involved in the biosynthesis of oligosaccharide chains.
 THERE ARE THREE MAJOR CLASSES OF GLYCOPROTEINS
 Glycoproteins can be divided into three main groups, based on the nature of the
linkage between the polypeptide and oligosaccharide chains:

an O-linkage (N-acetylgalactosamine to serine)

an N-linkage (Nacetylglucosamine
to asparagine) a glycosylphosphatidylinositol
(GPI) linkage

 The GPI structure shown is that linking acetylcholinesterase to the plasma

membrane of the human red blood cell.
o The site of action of PI-phospholipase C (PI-PLC), which releases the enzyme
from membrane binding is indicated.
 o This particular GPI contains an extra fatty acid attached to inositol and also
an extra phosphoryl-ethanolamine moiety attached to the central of the
mannose residue.

1. Those containing an O-glycosidic linkage (O-linked), involving the hydroxyl side

chain of serine or threonine (and sometimes also tyrosine) and a sugar such as N-
acetylgalactosamine (GalNAcSer[Thr]).
2. Those containing an N-glycosidic linkage (N-linked), involving the amide nitrogen of
asparagine and N-acetylglucosamine (GlcNAc-Asn).
3. Those linked to the carboxyl-terminal amino acid of a protein via a
phosphorylethanolamine moiety joined to an oligosaccharide (glycan), which in turn
is linked via glucosamine to phosphatidylinositol (PI).
a These are glycosylphosphatidylinositol-anchored (GPI-anchored)
glycoproteins. Among other functions, they are involved in directing
glycoproteins to the apical or basolateral areas of the plasma membrane of
polarized epithelial cells.
 The number of oligosaccharide chains attached to one protein can vary from 1 to
30 or more, with the sugar chains ranging from one or two residues in length to
much larger structures.
 The glycan chain may be linear or branched.
 Many proteins contain more than one type of sugar chain;
o for instance, glycophorin, an important red cell membrane glycoprotein,
contains both O- and N-linked oligosaccharides.
GLYCOPROTEINS CONTAIN SEVERAL TYPES OF O-GLYCOSIDIC
LINKAGES
At least four subclasses of O-glycosidic linkages are found in human glycoproteins:
1. The N-acetylgalactosamine-Ser(Thr) linkage shown in Figure 46–1 is the
predominant linkage. Usually a galactose or an Nacetylneuraminic acid residue is
attached to the N-acetylgalactosamine, but many variations in the sugar
compositions and lengths of such oligosaccharide chains are found. This type of
linkage is found in mucins (see below).
2. Proteoglycans contain a galactose-galactose-xylose trisaccharide (the so-called link
trisaccharide) attached to serine.
3. Collagens (see Chapter 50) contain a galactose-hydroxylysine linkage.
4. Many nuclear and cytosolic proteins contain side chains consisting of a single N-
acetylglucosamine attached to a serine or threonine residue.
Mucins Have a High Content of O-Linked Oligosaccharides &
Exhibit Repeating Amino Acid Sequences

Mucins
 are glycoproteins that are highly resistant to proteolysis because the density of
oligosaccharide chains makes it difficult for proteases to access the polypeptide
chain.
 They help to lubricate and form a protective physical barrier on epithelial
surfaces.
 Two distinctive characteristics:
o high content of O-linked oligosaccharides (the carbohydrate content is
generally more than 50%);
o presence of variable numbers of tandem repeats (VNTRs) of peptide
sequence in the center of the polypeptide chain, to which the Oglycan
chains are attached in clusters.
 These tandem repeat sequences are rich in serine, threonine, and
proline; indeed, up to 60% of the dietary requirement for threonine
can be accounted for by the synthesis of mucins.
o also contain a number of N-glycan chains.
 Both secretory and membrane-bound mucins occur:
 Secretory mucins:
o Mucus secreted by the gastrointestinal, respiratory, and reproductive tracts
is a solution containing about 5% mucins.
o Secretory mucins generally have an oligomeric structure, with monomers
linked by disulfide bonds, and hence a very high molecular mass.
o Mucus has a high viscosity and often forms a gel because of its content of
mucins.
o The high content of O-glycans confers an extended structure. This is partly
explained by steric interactions between the N-acetylgalactosamine
moieties and adjacent amino acids, resulting in a chain-stiffening effect, so
that the conformation of mucins often become rigid rods.
o Intermolecular noncovalent interactions between sugars on neighboring
glycan chains contribute to gel formation.
o The high content of N-acetylneuraminic acid and sulfate residues found in
many mucins gives them a negative charge.
 Membrane-bound mucins participate in cell–cell interactions, and may also mask
cell surface antigens.
  Many cancer cells form large amounts of mucins that mask surface antigens and
protect the cancer cells from immune surveillance.
 Mucins also carry cancer-specific peptide and carbohydrate epitopes. Some of
these have been used to stimulate an immune response against cancer cells.

O-Linked Glycoproteins Are Synthesized by Sequential Addition of

Sugars from Sugar Nucleotides

 Because most glycoproteins are membrane-bound or secreted, their mRNA is

usually translated on membrane-bound polyribosomes.
 The glycan chains are built up by the sequential donation of sugars from sugar
nucleotides, catalyzed by glycoprotein glycosyltransferases.
 Carrier systems (permeases and transporters) are needed to transport sugar
nucleotides (UDP-galactose, GDP-mannose, and CMP-Nacetylneuraminic acid)
across the Golgi membrane.
o They are antiporter systems; the influx of one molecule of sugar nucleotide
is balanced by the efflux of one molecule of the corresponding nucleotide
(UMP, GMP, or CMP).

 There are 41 different types of glycoprotein glycosyltransferases.

o Families of glycosyltransferases are named for the sugar nucleotide donor,
and subfamilies on the basis of the linkage formed between the sugar and
the acceptor substrate.
o transfer may occur with retention or inversion of the conformation at C-1 of
the sugar.
o Binding of the sugar nucleotide to the enzyme causes a conformational
change in the enzyme that permits binding of the acceptor substrate.
o Glycosyltransferases show a high degree of specificity for the acceptor
substrate; typically acting only on the product of the preceding reaction.
o The different stages in glycan formation, and hence the different
glycosyltransferases, are located in different regions of the Golgi, so that
there is spatial separation of the steps in the process.
o Not all of the glycan chains of a given glycoprotein are complete; some are
truncated, leading to microheterogeneity.
o It is either the serine and threonine residues are glycosylated, and the first
sugar moiety incorporated is usually N-acetylgalactosamine.
N-LINKED GLYCOPROTEINS CONTAIN AN ASPARAGINE-N-
ACETYLGLUCOSAMINE LINKAGE

 N-Linked glycoproteins are the major class of glycoproteins, including both

membrane-bound and circulating glycoproteins.
o They are distinguished by the presence of the asparagine—N-
acetylglucosamine linkage.
o There are three major classes of N-linked oligosaccharides: complex, high-
mannose, and hybrid. All three classes have the same branched
pentasaccharide, Man3GlcNAc2, bound to asparagine, but differ in their
outer branches.

Structures of the major types of asparagine-linked oligosaccharides.

 The boxed area encloses the pentasaccharide core common to all N-linked
glycoproteins.
 Complex oligosaccharides contain two, three, four, or five outer branches.
 The oligosaccharide branches are often referred to as antennae, so that bi-, tri-, tetra-,
and penta-antennary structures may all be found.
  They generally contain terminal N-acetylneuraminic acid residues and underlying
galactose and N-acetylglucosamine residues, the latter often constituting the
disaccharide N-acetyllactosamine

The Biosynthesis of N-Linked Glycoproteins Involves Dolichol-P-P-

Oligosaccharide
 The synthesis of all N-linked glycoproteins
o begins with the synthesis of a branched oligosaccharide attached to dolichol
pyrophosphate on the cytosolic side of the endoplasmic reticulum
membrane

The structure of dolichol

o then translocated to the lumen of the endoplasmic reticulum, where it

undergoes further glycosylation
o before the oligosaccharide chain is transferred by an oligosaccharyl
transferase onto an asparagine residue of the acceptor apoglycoprotein as it
enters the endoplasmic reticulum during synthesis on membrane-bound
polyribosomes.
 Pathway of biosynthesis of dolichol pyrophosphate
oligosaccharide.

As shown in the pathway:

 the first step is a reaction between UDP-Nacetylglucosamine
and dolichol phosphate, forming N-acetylglucosamine dolichol
pyrophosphate.
 A second N-acetylglucosamine is added from UDP-N-
acetylglucosamine, followed by the addition of five molecules
of mannose from GDP-mannose.
 The dolichol pyrophosphate oligosaccharide is then
translocated into the lumen of the endoplasmic reticulum,
and further mannose and glucose molecules are added, to
form the final dolichol pyrophosphate oligosaccharide, using
dolichol phosphate mannose and dolichol phosphate glucose
as the donors. The dolichol pyrophosphate oligosaccharide is
then transferred onto the acceptor asparagine residue of the
nascent protein chain.

 To form high-mannose chains, the glucose and some of the

peripheral mannose residues are removed by glycosidases.

 To form an oligosaccharide chain of the complex type, the

glucose residues and four of the mannose residues are
removed by glycosidases in the endoplasmic reticulum and
Golgi, then N-acetylglucosamine, galactose, and
Nacetylneuraminic acid are added in reactions catalyzed by
glycosyltransferases in the Golgi apparatus.
 Hybrid chains are formed by partial processing, forming complex chains on
one arm and mannose units on the other arm.
Glycoproteins & Calnexin Ensure Correct Folding of Proteins in the
Endoplasmic Reticulum

Calnexin. (a) Overall structure of calnexin showing the globular domain with a legume lectin-like β-sandwich fold and the
extended P-domain thought to embrace the glycoprotein substrate (PDB 1JHN). (b) Closeup view of the carbohydrate-
binding binding site with a glucose molecule (green) modeled into it according to the description in reference 35. Selected
residues are labeled.

 Calnexin is a chaperone protein in the endoplasmic reticulum membrane; binding

to calnexin prevents a glycoprotein from aggregating.
o It is a lectin, recognizing specific carbohydrate sequences in the glycan chain
of the glycoprotein.
o Incorrectly folded glycoproteins undergo partial deglycosylation, and are
targeted to undergo transport from the endoplasmic reticulum back to the
cytosol for catabolism.
o Calnexin binds to glycoproteins that possess a monoglycosylated core from
which the terminal glucose residue has been removed, leaving only the
innermost glucose attached.
o Calnexin and the bound glycoprotein form a complex with ERp57, a homolog
of protein disulfide isomerase, which catalyzes disulfide bond interchange,
facilitating correct folding.
o The bound glycoprotein is released from its complex with calnexin-ERp57
when the sole remaining glucose is hydrolyzed by a glucosidase and is then
available for secretion if it is correctly folded.
 If not correctly folded, a glucosyltransferase recognizes this and
reglucosylates the glycoprotein, which rebinds to the calnexin-Erp57
complex.
 If it is now correctly folded, the glycoprotein is again deglucosylated
and secreted.
 If it is not capable of correct folding, it is translocated out of the
endoplasmic reticulum into the cytosol for catabolism.
 The glucosyltransferase senses the folding of the glycoprotein and only
reglucosylates misfolded proteins.
 The soluble endoplasmic reticulum protein calreticulin performs a
similar function to that of calnexin.
Several Factors Regulate the Glycosylation of Glycoproteins
 The glycosylation of glycoproteins involves a large number of enzymes; about 1%
of the human genome codes for genes that are involved with protein
glycosylation. There are at least 10 distinct N-acetylglucosamine transferases, and
multiple isoenzymes of the other glycosyltransferases. Controlling factors in the
first stage of N-linked glycoprotein biosynthesis (assembly and transfer of the
dolichol pyrophosphate oligosaccharide) include not only the availability of the
sugar nucleotides, but also the presence of suitable acceptor sites in proteins, the
tissue concentration of dolichol phosphate, and the activity of the oligosaccharide:
protein transferase.

 Various cancer cells are found to synthesize different oligosaccharide chains from
those made in normal cells (eg, greater branching). This is due to cancer cells
expressing different patterns of glycosyltransferases from those in normal cells, as
a result of specific gene activation or repression. The differences in
oligosaccharide chains could affect adhesive interactions between cancer cells and
the normal parent tissue cells, contributing to metastasis.
SOME PROTEINS ARE ANCHORED TO THE PLASMA MEMBRANE BY
GLYCOPHOSPHATIDYL-INOSITOL STRUCTURES

 Membrane-bound proteins that are anchored to the lipid bilayer by a

glycophosphatidylinositol (GPI) tail (Figure 46–1) constitute the third major class
of glycoproteins. GPI linkage is the commonest way in which various proteins are
anchored to cell membranes.
 The proteins are anchored to the outer leaflet of the plasma membrane or the
inner (luminal) leaflet of the membrane in secretory vesicles by the fatty acids of
phosphatidylinositol. The phosphatidylinositol is linked via N-acetylglucosamine to
a glycan chain containing a variety of sugars, including mannose and glucosamine.
In turn, the oligosaccharide chain is linked via phosphorylethanolamine in an
amide linkage to the carboxylterminal amino acid of the protein. Additional
constituents are found in many GPI structures; for example, that shown in Figure
46–1 contains an extra phosphorylethanolamine attached to the middle of the
three mannose moieties of the glycan and an extra fatty acid attached to
glucosamine.

There are three functions of this GPI linkage:

1. The GPI anchor allows greatly enhanced mobility of a protein in the plasma
membrane compared with that of a protein that contains transmembrane
sequences. The GPI anchor is attached only to the outer leaflet of the lipid bilayer, so
that it is freer to diffuse than a protein anchored through both layers of the
 membrane. Increased mobility may be important in facilitating rapid responses to
stimuli.
2. Some GPI anchors may connect with signal transduction pathways, so that proteins
that do not have a transmembrane domain may nevertheless be receptors for
hormones and other cell surface signals.
3. GPI structures can target proteins to apical or basolateral domains of the plasma
membrane of polarized epithelial cells.

 The GPI anchor is preformed in the endoplasmic reticulum, and is then attached to
the protein after ribosomal synthesis is complete.
 The primary translation products of GPI-anchored proteins have not only an
amino terminal signal sequence that directs them into the endoplasmic reticulum
during synthesis, but also a carboxy-terminal hydrophobic domain that acts as the
signal for attachment of the GPI anchor.
 The first stage in synthesis of the GPI anchor is insertion of the fatty acids of
phosphatidylinositol into the luminal face of the endoplasmic reticulum
membrane, followed by glycosylation, starting with esterification of N-acetyl-
glucosamine to the phosphate group of phosphatidylinositol.
 A terminal phosphoethanolamine moiety is added to the completed glycan chain.
The hydrophobic carboxyterminal domain of the protein is displaced by the amino
group of ethanolamine in the transamidation reaction that forms the amide
linkage between the GPI anchor and an aspartate residue in the protein.
SOME PROTEINS UNDERGO RAPIDLY REVERSIBLE GLYCOSYLATION
 Many proteins, including nuclear pore proteins, proteins of the cytoskeleton,
transcription factors, and proteins associated with chromatin, as well as nuclear
oncogene proteins and tumor suppressor proteins, undergo rapidly reversible O-
glycosylation with a single sugar moiety, Nacetylglucosamine.
 The serine and threonine sites of glycosylation are the same as those of
phosphorylation of these proteins, and glycosylation and phosphorylation occur
reciprocally in response to cellular signaling.
 The O-linked N-acetylglucosamine transferase that catalyzes this glycosylation
uses UDP-N-acetylglucosamine as the sugar donor, and has phosphatase activity,
so can directly replace a serine or threonine phosphate with N-acetylglucosamine.
There is no absolute consensus sequence for the reaction, but about half the sites
that are subject to reciprocal glycosylation and phosphorylation are Pro-Val-Ser.
O-linked Nacetylglucosamine transferase is activated by phosphorylation in
response to insulin action, and the N-acetylglucosamine is removed by
Nacetylglucosaminidase, leaving the site available for phosphorylation. Both the
activity and peptide specificity of O-linked Nacetylglucosamine transferase depend
on the concentration of UDP-Nacetylglucosamine. Depending on cell type, up to 2
to 5% of glucose metabolism is by way of the hexosamine pathway leading to
Nacetylglucosamine formation, giving the O-linked N-acetylglucosamine
transferase a role in nutrient sensing in the cell.
 Excessive O-glycosylation with N-acetylglucosamine (and hence reduced
phosphorylation) of target proteins is implicated in insulin resistance and glucose
toxicity in diabetes mellitus, as well as neurodegenerative diseases.
ADVANCED GLYCATION END-PRODUCTS (AGEs) ARE IMPORTANT
IN CAUSING TISSUE DAMAGE IN DIABETES MELLITUS
Glycation is the nonenzymic attachment of sugars (mainly glucose) to amino groups of
proteins (and also to other molecules including DNA and lipids), unlike glycosylation
which is enzyme-catalyzed. Initially, glucose forms a Schiff base to the amino terminal of
the protein, which then undergoes the Amadori rearrangement to yield ketoamines
(Figure 46– 5), and further reactions to yield advanced glycation end-products (AGEs).
The overall series of reactions is known as the Maillard reaction, which is involved in the
browning of certain foodstuffs during storage or heating, and provides much of the
flavor of some foods.

Formation of advanced glycation end-products from glucose.

Advanced glycation end-products underlie tissue damage in poorly controlled

diabetes mellitus. When the blood glucose concentration is consistently elevated, there
is increased glycation of proteins. Glycation of collagen and other proteins in the
extracellular matrix alters their properties (eg, increasing the cross-linking of collagen).
Cross-linking can lead to accumulation of various plasma proteins in the walls of blood
vessels; in particular, accumulation of low-density lipoprotein (LDL) can contribute to
atherogenesis. AGEs appear to be involved in both microvascular and macrovascular
damage in diabetes mellitus. Endothelial cells and macrophages have AGE receptors on
their surfaces; uptake of glycated proteins by these receptors can activate the
transcription factor NF-kB, generating a variety of cytokines and proinflammatory
molecules.
Nonenzymic glycation of hemoglobin A present in red blood cells leads to the
formation of HbA1c. It occurs normally to a small extent, and is increased in diabetic
patients with poor glycemic control, whose blood glucose concentration is consistently
elevated. Measurement of HbA1c has become a very important part of the management
of patients with diabetes mellitus.
GLYCOPROTEINS ARE INVOLVED IN MANY BIOLOGICAL PROCESSES
& IN MANY DISEASES
Glycoproteins have many different functions, including transport molecules,
immunological molecules, and hormones. They are also important in fertilization and
inflammation; a number of diseases are due to defects in the synthesis and catabolism
of glycoproteins.

Glycoproteins Are Important in Fertilization

 To reach the plasma membrane of an oocyte, a sperm has to traverse the zona
pellucida (ZP), a thick, transparent, envelope that surrounds the oocyte. The
glycoprotein ZP3 is an O-linked glycoprotein that functions as a sperm receptor. A
protein on the sperm surface interacts with the oligosaccharide chains of ZP3. By
transmembrane signaling, this interaction induces the acrosomal reaction, in
which enzymes such as proteases and hyaluronidase and other contents of the
acrosome of the sperm are released. Liberation of these enzymes permits the
sperm to pass through the zona pellucida and reach the plasma membrane of the
oocyte. Another glycoprotein, PH-30, is important in binding of the sperm plasma
membrane to that of the oocyte, and the subsequent fusion of the two
membranes, enabling the sperm to enter and fertilize the oocyte.

Selectins Play Key Roles in Inflammation & in Lymphocyte Homing

 Leukocytes play important roles in many inflammatory and immunological
processes; the first steps are interactions between circulating leukocytes and
endothelial cells prior to passage of the leukocytes out of the circulation.
Leukocytes and endothelial cells contain cell surface lectins, called selectins, which
2+
participate in intercellular adhesion. Selectins are single-chain Ca -binding
transmembrane proteins; the amino terminals contain the lectin domain, which is
involved in binding to specific carbohydrate ligands.
 Interactions between selectins on the neutrophil cell surface and glycoproteins on
the endothelial cell trap the neutrophils temporarily, so that they roll over the
endothelial surface. During this the neutrophils are activated, undergo a change in
shape, and now adhere firmly to the endothelium. This adhesion is the result of
interactions between integrins on the neutrophils and immunoglobulin-related
proteins on the endothelial cells. After adhesion, the neutrophils insert
pseudopodia into the junctions between endothelial cells, squeeze through these
 junctions, cross the basement membrane, and are then free to migrate in the
extravascular space.
 Selectins bind sialylated and fucosylated oligosaccharides. Sulfated lipids may
also be ligands. Synthesis of compounds such as monoclonal antibodies that block
selectin-ligand interactions may be therapeutically useful to inhibit inflammatory
responses. Cancer cells often have selectin ligands on their surfaces, which may
have a role in the invasion and metastasis of cancer cells.

Abnormalities in the Synthesis of Glycoproteins Underlie Certain

Diseases

Examples:
 Leukocyte adhesion deficiency II is a rare condition due to mutations affecting the
activity of a Golgi-located GDP-fucose transporter. The absence of fucosylated
ligands for selectins leads to a marked decrease in neutrophil rolling. Patients
suffer life-threatening recurrent bacterial infections, and also psychomotor and
mental retardation. The condition may respond to oral fucose.

 Paroxysmal nocturnal hemoglobinuria is an acquired mild anemia characterized

by the presence of hemoglobin in urine due to hemolysis of red cells, particularly
during sleep, reflecting a slight drop in plasma pH during sleep, which increases
susceptibility to lysis by the complement system. The condition is due to the
acquisition in hematopoietic cells of somatic mutations in the gene coding for the
enzyme that links glucosamine to phosphatidylinositol in the GPI structure. This
leads to a deficiency of proteins that are anchored to the red cell membrane by
GPI linkage. Two proteins, decay accelerating factor and CD59 normally interact
with components of the complement system to prevent the hemolysis. When they
are deficient, the complement system acts on the red cell membrane to cause
hemolysis.

 Some of the congenital muscular dystrophies are the result of defects in the
synthesis of glycans in the protein α-dystroglycan. This protein protrudes from the
surface membrane of muscle cells and interacts with laminin-2 (merosin) in the
basal lamina. If the glycans of α-dystroglycan are not correctly formed (as a result
 of mutations in genes encoding some glycosyltransferases), this results in
defective interaction of α-DG with laminin.

 Rheumatoid arthritis is associated with an alteration in the glycosylation of

circulating immunoglobulin G (IgG) molecules, such that they lack galactose in
their Fc regions and terminate in N-acetylglucosamine. Mannose-binding protein,
a lectin synthesized by liver cells and secreted into the circulation, binds mannose,
Nacetylglucosamine, and some other sugars. It can thus bind agalactosyl IgG
molecules, which subsequently activate the complement system, contributing to
chronic inflammation in the synovial membranes of joints. Mannose-binding
protein can also bind sugars when they are present on the surfaces of bacteria,
fungi, and viruses, preparing these pathogens for opsonization or for destruction
by the complement system. This is an example of innate immunity, not involving
immunoglobulins or T lymphocytes. Deficiency of this protein in young infants as a
result of mutation renders them susceptible to recurrent infections.
Inclusion Cell (I-Cell) Disease Results From Faulty Targeting of
Lysosomal Enzymes
 Mannose 6-phosphate serves to target enzymes into the lysosome.
 I-cell disease is a rare condition characterized by severe progressive psychomotor
retardation and a variety of physical signs, with death often occurring in the first
decade of life.
o Cells from patients with I-cell disease lack almost all of the normal lysosomal
enzymes; the lysosomes thus accumulate many different types of
undegraded molecules, forming inclusion bodies.
o The patients’ plasma contains very high activities of lysosomal enzymes,
suggesting that the enzymes are synthesized but fail to reach their proper
intracellular destination and are instead secreted.
 Lysosomal enzymes from normal individuals carry the mannose-6-phosphate
recognition marker
 Cells from patients with I-cell disease lack the Golgi-located N-acetylglucosamine
phosphotransferase.
 Two lectins act as mannose 6-phosphate receptor proteins;
o both function in the intracellular sorting of lysosomal enzymes into clathrin-
coated vesicles in the Golgi—these vesicles then leave the Golgi and fuse
with a prelysosomal compartment.

Genetic Deficiencies of Glycoprotein Lysosomal Hydrolases Cause

Diseases Such as α-Mannosidosis
 Turnover of glycoproteins involves catabolism of the oligosaccharide chains
catalyzed by a number of lysosomal hydrolases, that includes:
o α-neuraminidase
o β-galactosidase
o β-hexosaminidase
o α- and βmannosidases
o α-N-acetylgalactosaminidase
o α-fucosidase
o endo-β-Nacetylglucosaminidase
o aspartylglucosaminidase.
 Genetic defects of these enzymes result in abnormal degradation of glycoproteins.
 o The accumulation in tissues of partially degraded glycoproteins leads to
various diseases, including:
 Mannosidosis
 Fucosidosis
 Sialidosis
 Aspartylglucosaminuria
 Schindler disease, due respectively to deficiencies of α-mannosidase,
α-fucosidase, αneuraminidase, aspartylglucosaminidase, and α-
Nacetylgalactosaminidase.

GLYCANS ARE INVOLVED IN THE BINDING OF VIRUSES, BACTERIA,

& SOME PARASITES TO HUMAN CELLS
feature of glycans:
 they bind specifically to proteins and other glycans
o ability to bind some viruses, bacteria, and parasites.

Examples:

- Influenza virus A binds to cell surface glycoprotein receptor molecules containing N-

acetylneuraminic acid via a hemagglutinin protein.
o The virus also has a neuraminidase that plays a key role in allowing elution of
newly synthesized progeny from infected cells. If this process is inhibited,
spread of the viruses is markedly diminished.
 Inhibitors of this enzyme (eg, zanamivir, oseltamivir) are now available
for use in treating patients with influenza.
o Influenza viruses are classified according to the type of hemagglutinin (H) and
neuraminidase (N) that they possess.
 There are at least 16 types of hemagglutinin and 9 types of
neuraminidase.
 avian influenza virus is classified as H5N1.

- Human immunodeficiency virus type 1 (HIV-1), the cause of AIDS, attaches to cells
via one of its surface glycoproteins (gp 120) and uses another surface glycoprotein
(gp 41) to fuse with the host cell membrane.
 o Antibodies to gp 120 develop during infection by HIV-1, and there has been
interest in using the protein as a vaccine.
 One major problem with this approach is that the structure of gp 120 can
change relatively rapidly due to mutations, allowing the virus to escape
from the neutralizing activity of antibodies directed against it.

- Helicobacter pylori is the major cause of peptic ulcers.

o It binds to at least two different glycans present on the surfaces of epithelial
cells in the stomach allowing it to establish a stable attachment site to the
stomach lining.

- Many bacteria that cause diarrhea attach to surface cells of the intestinal mucosa via
glycans present in glycoproteins or glycolipids.
o The attachment of the malarial parasite Plasmodium falciparum to human cells
is mediated by a GPI present on the surface of the parasite.