Faculty of Biochemistry and Molecular Medicine
Biomolecules for Biochemists (8 op)
740147P Biomolecules for Bioscientists (8 op)
740148P Biomolecules (5 op)
740143P
CARBOHYDRATES
Docent Tuomo Glumoff
1
CARBOHYDRATES lectures
Figures under © taken from the following sources:
Horton et al., Principles of Biochemistry
Mathews et al., Biochemistry
Campbell et al., Biochemistry Illustrated
Cooper & Hausman: The Cell – A Molecular Approach
Devlin: Textbook of Biochemistry with Clinical Correlations
Nelson & Cox, Lehninger Principles of Biochemistry
Voet & Voet, Biochemistry
Berg et al., Biochemistry
Varki et al. (ed.) Essentials of Glycobiology
Rawn: Biochemistry
Recommended text books for further reading – any standard biochemistry text book, and certainly one of the following:
Voet et al., Principles of Biochemistry, 3rd edition (2008), chapter 8 (Wiley ISBN-13: 978-0470-23396-2)
Mathews et al., Biochemistry, 4th edition (2013), chapter 9 (Pearson ISBN 978-0-13-800464-4)
Answers to problems will be given at the lectures and they will also be available after the lectures in NOPPA.
1. Basics of carbohydrates
- carbohydrates = saccharides ≈ sugars
- What is a sugar? How to define a sugar?
Typical properties of sugars?
• sweet taste
• crystallizes
• water-soluble
• polyhydroxy aldehydes or
polyhydroxy ketones
• contains a chiral carbon
• optically active (stereoisomers)
• for monosaccharides the molecular
formula is (CH2O)n, where 3  n  9
H
H
O
H
O
H
H
O
O
H
OH
H
H
H
O
O
O
H
H
O
O
O
H
O
H
H
O
H
H
H
H
O
O
H
*)
H
H
H
(CH2O)n is an ”oversimplification”, since many saccharides are modified and some contain
also atoms such as S and N. Nevertheless, all compounds herewith either have this formula
or can be derived from substances that do. (Mathews et al. Biochemistry 4th edition)
PROBLEM 1. Why a 2-carbon compound cannot be a sugar although it would fulfill the formula
(CH2O)n?
- most abundant class of biological
molecules on earth by mass
- all organisms can synthesize, but
most important is photosynthesis:
solar energy to chemical energy
- mono- and disaccharides are
usually metabolites
- oligosaccharides are usually linked
to proteins or lipids
- polysaccharides are usually storage
forms or have structural function
2. Monosaccharides
2
Monosaccharides
- trioses, 3 carbons
- ketoses and aldoses are tautomers
(an aldotriose)
(a ketotriose)
PROBLEM 2. Please check that the formula (CH2O)n hold for both triose tautomers!
- D- and L-enantiomers = isomers differing at the chiral carbon configuration
• mirror images
- in nature D-monosaccharides dominate
- D-L vs. R-S naming system:
• R- and S- would be absolute, but becomes difficult with multiple chiral carbons
• +/- and d/l (to right vs. to left) could be used with respect to ability to rotate planepolarized light
PROBLEM 3. You will be given molecular models of an R-isomer and an S-isomer. Which is which?
red =
blue =
3
- tetroses, 4 carbons
- may have 2 chiral carbons, and thus are diastereomers
- D- or L- is assigned based on the chiral carbon farthest from the carbonyl group (= highest
numbered chiral C) and additional names are given for diastereomers (an arbitrary naming system)
- epimers = multiple D/L possibilities, but a difference in only one of them
- pentoses, 5 carbons
• for example riboses in nucleotides
- hexoses, 6 carbons
• typical energy molecules, like glucose
4
D-sugars have the same absolute
configuration at the asymmetric center
furthest from their carbonyl group as does
D-glyceraldehyde
Examples of monosaccharides with their occurrence and physiological role:
PROBLEM 4. Observe the aldose and ketose structures on page 5 and answer the following questions:
4.1 Why are there a fewer number of different ketoses?
4.2 Identify each of the following:
a) two aldoses whose configuration at carbons 3, 4 and 5 matches that of D-fructose
b) the enantiomer of D-galactose
c) an epimer of D-galactose that is also an epimer of D-mannose
d) a ketose that has no chiral centers
e) a ketose that has one chiral center
Fischer projections of C3 to C6 aldoses
5
carbon
numbering:
1
2
3
4
Fischer projections of C3 to C6 ketoses
- the open chain form of a monosaccharide is in equilibrium with the ring form
- under physiological conditions ≥ 5-carbon monosaccharides are ≥ 99% in the ring form
6
Pyranose formation:
The open-chain form of glucose
cyclizes when the C-5 hydroxyl
group attacks the oxygen atom
of the C-1 aldehyde group to
form an intramolecular
hemiacetal. Two anomers,
designated a and b, can result.
- Haworth projection: thick edge
towards the viewer, thin edge
towards the rear
- interconversion
between a and b forms
= mutarotation
- catalyzed by specific
mutarotases
trans
a = on the opposite side
cis
b = on the same side
7
6
1
5
2
4
3
- hemiacetal or hemiketal bond is formed
- pentoses and hexoses form pyran and
furan rings => pyranoses and
furanoses
- anomers = upon cyclization the former
carbonyl carbon becomes asymmetric =>
anomeric carbon
- note that pentoses and hexoses may both form 5- and 6-rings (size of ring not dependent on
number of carbons!)
- distribution between furanose and pyranose depends on the sugar, pH, solvent and temperature
*
* glucose: open chain form 0.2 %
8
PROBLEM 5. On page 10 you will find the
Haworth projection of b-D-Nacetylglucosamine. Draw it using the
Fischer projection (open-chain form).
PROBLEM 6. Take mannose from the tables
on page 5 and draw it in a ring form.
chair and boat conformations:
- same stereochemistry, difference in three-dimensional shape
- conformations with bulky substrates in equatorial positions are favored, while steric hindrance of
axial substituents renders the boat conformation less favored
e = equatorial
a = axial
- conformations: interchangeable by simple deformation of the molecule
- configurations: interchangeable only by breaking and reformation of covalent bonds
9
- monosaccharides may contain substituents or modifications that make them derivatives
- examples:
- aminosugars:
- reduction of the carbonyl
group => alditol
(xylitol is accordingly made
from xylose)
glyceraldehyde-3-phosphate
glucose-1-phosphate
glucose-6-phosphate
fructose-6-phosphate
- sugar phosphates are very
important derivatives
- activated molecules in energy
metabolism
10
- glycosides:
a glycosidic bond is formed
11
- glycosidic bond is the bond
between two sugar units in
di- and polysaccharides,
but as seen here the
glycosidic bond can also
form between a sugar and
another group
- glycosidic bond is also
present in nucleotides in
DNA and RNA
- common abbreviations used for monosaccharides
12
13
3. Disaccharides
- two monosaccharides are joined with a glycosidic bond
=> hydroxyl group of one sugar reacts with the hydroxyl group of the anomeric carbon of another sugar
NOTE!!
- equilibrium to
left
- polysaccharides
are metastable
compounds
- hemiacetal becomes acetal
- in common disaccharides O-glycosidic bond; in glycoproteins and nucleotides N-glycosidic bond
- glycosidic bond is readily hydrolyzed in acid, but resists bases
- oxidation of the anomeric carbon means that the sugar is a reducing sugar (remember:
oxidation and reduction are always coupled – the compound that gets oxidized will leave
another compound reduced)
- a reducing sugar has a reducing end and a nonreducing end
- reducing end is the one with a free anomeric carbon (and thus also a free aldehyde group)
- only the open-chain form can undergo oxidation, not the hemiacetal (ring) form
- in a disaccharide only the rightmost monomer can adopt the open chain form
14
- mutarotation may also take place in disaccharides, namely the reducing end monosaccharide
- a,a form is simply “a”, while “a,b” is simply b
- if the disaccharide ends in a nonreducing end, it cannot mutarotate (such as sucrose)
Here is an a-disaccharide. Please draw the corresponding b-disaccharide.
PROBLEM 7. One of the disaccharides on the following page is not a reducing sugar? Which
one and why?
Distinguishing features of different disaccharides:
15
1. The two monomers may be of the same kind, or
they may be different
2. The most common linkages between two
monosaccharides are:
11 12 14 16
(at least one anomeric hydroxyl is always involved
in the bond)
3. The order of the two monomers (if different)
determines if the disaccharide can be a reducing
sugar or not.
4. The anomeric configuration of the hydroxyl group
on carbon 1 of each monomer determine which
enzyme can hydrolyze the glycosidic bond
(maltose and cellobiose are both made up from
two glucoses, but cannot serve as a substrate for
the same hydrolytic enzyme) - Why?
Disaccharides in 3D for example from here:
www.biotopics.co.uk/JmolApplet/maltosejdisplay.htm
Compare maltose with cellobiose!
Nomenclature of disaccharides (systematic names):
Reducing disaccharides (contain a free
hemiacetal):
"glycosylglycose"
example:
α-D-Glucopyranosyl-(1  4)-β-D-glucopyranose
(trivial name β-maltose)
Non-reducing disaccharides (without a free
hemiacetal):
"glycosyl glycoside"
example:
α-D-Glucopyranosyl- (1  1)- α-D-glucopyranoside
(trivial name α,α-trehalose)
16
Names and writing of disaccharides:
- sequence starts from the left
(nonreducing end)
- anomeric and enantiomeric forms can be
designated
- the ring configuration can be given
- the atoms between which the glycosidic
bond is formed are given
(the arrow shows the direction from the
anomeric carbon)
maltose:
a-D-Glcp(14)-b-D-Glcp
or
sucrose:
Glc(a1b4)Glc
a-D-Glcp(1↔2)-b-D-Fruf
"invert sugar" = hydrolysis of sucrose to glucose
and fructose
PROBLEM 8. Write below a systematic name for lactose.
4. Polysaccharides
= glycans
- storage polysaccharides like, starch and glycogen, as well as stability, biosynthesis and degradation
of polysaccharides will be postponed to Aineenvaihdunta I (Metabolism I) course later in the spring
- here we will study other types of oligo- and polysaccharides and their functions
- homopolysaccharides – repeating one type of units
- heteropolysaccharides – repeating usually two kinds of units
- complex polysaccharides – contain more than two kinds of units
- structural elements in plant cell walls and animal exoskeletons
- extracellular support – bacterial cell envelope and in animals a matrix that holds cells together and
supports tissues and organs
17
- no defined molecular weight = saccharide chains may grow to become “shorter or longer”
(But NOTE: saccharide chains with specific function e.g. in recognition must be of certain length and
composition; see later blood group antigens)
18
Cellulose:
- linear polymer of b-D-glucose, 10.000
to 15.000 units
- most abundant polysaccharide
molecule on earth
Chitin:
- linear polymer of b-N-acetylglucosamine
- found in surface armor of insects
Glycosaminoglycans:
- polymers of repeating disaccharide units
- one of the sugars is either N-acetylgalactosamine or N-acetylglucosamine or a derivative thereof
- are acids through either a sulfate or carboxylate group present
- examples of modified sugar residues that gain novel properties and functions
- heparin is a natural anticoagulant in body fluids
(here only the repeating unit shown!)
- inhibits blood clotting enzymes through binding
to anti-prothrombin III protein
19
GlcUA
GlcUA
- hyaluronic acid is noted for its much longer chain than most other glycosaminoglycans
PROBLEM 9. Chain length of a polysaccharide (number of units in a chain) is not known in
the beginning of its synthesis, i.e. it is not predefined. See p. 19: the numbers are not precise
– sometimes the glycan chain is “shorter”, sometimes “longer”. Please explain!
20
Glycoconjugates: proteoglycans, glycoproteins and glycolipids
- proteoglycan is a protein-carbohydrate complex found in extracellular space
- peptidoglycan is an important part of bacterial cell walls
Left: Gram negative bacilli
Right: Gram positive cocci
21
22
- for comparison, the cell wall of mycobacteria is even more complicated in structure (figure below)
- on top of the peptidoglycan layer there are arabinogalactan (carbohydrate) and mycolic acid
(lipid) layers
- this makes the cell wall more resistant and contributes to the difficulty of eliminating tuberculosis
causing mycobacteria
23
24
galectin = galactoside-binding lectin
lectin = sugar-binding protein
- an example, how membrane proteins and carbohydrate structures attached to them form part of
the cell´s contact network
- cell to cell
- cell to extracellular matrix
- protein-protein interactions
- protein-carbohydrate interactions
- cell surface structures mediated via carbohydrates can be part of intracellular signaling
25
Cooper & Hausman: The Cell
crosslinked
tetrapeptides
Voet et al.: Principles of Biochemistry
- glycoproteins are proteins that contain certain types of carbohydrate chains attached to them
- O-linked glycoproteins and N-linked glycoproteins
- N-linked glycans are attached to asparagine residues in a sequence –Asn – X – Ser/Thr –
- O-linked glycans are attached to threonines or serines
- glycan chains can be of various composition and type of branching
- glycans are attached to protein cotranslationally in the ER (endoplasmic reticulum)
- glycans are further modified in the Golgi apparatus
- glycan chains give the cellular machinery extra possibilities for recognizing proteins and cells
• immunoglobulin tissue distribution and interaction with phagocytic cells
• intracellular targeting and excretion
• cellular identification
• control of body fluid viscosity
• blood groups
- other functions of glycans include:
• stabilize protein fold (ready-made protein)
• stabilize protein folding (during the folding!) by binding to intermediate conformations
• stiffens and extends the polypeptide chain
26
- note that saccharide chains are actually surprisingly large in size compared to proteins
– a chain of a couple of sugar residues easily make a structure with some dimensions
comparable to a protein molecule
27
PROBLEM 10. E. coli is often used in the
lab to produce cloned eukaryotic proteins.
Sometimes the proteins cannot be obtained
in good amounts (there can be many
reasons). Can you identify or guess one
possible reason from the material we just
discussed?
- oligosaccharide chains attached to membrane proteins
at the surface of red blood cells
- to attach the terminal monosaccharide to get either
A or B blood group, one needs specific enzymes;
heterozygotes have both to get AB
- type O oligosaccharides are nonantigenic
PLEASE NOTE!!!!
(refer to problem 9.)
Whenever a glycan
chain has a specific
recognition function, the
number and the sugar
structure must be
precise!!!!!
- glycolipids and lipopolysaccharides are membrane components, where carbohydrates are
attached to lipids
28
Lipid layer
© W.W. Christie
The Scottish Crop Research Institute, Dundee
- lipopolysaccharides (LPS) can be found on the
surface of bacteria, like E. coli and Salmonella
- antibodies are raised in the body against LPS
to fight bacterial infections
- LPS of some bacteria are toxic
PROBLEM 11. Write down the distinctive differences of the cell wall structures between Gram-positive
and –negative bacteria.
Good to be aware of for possible further need:
Nomenclature of Carbohydrates:
http://www.chem.qmul.ac.uk/iupac/2carb/
INTERNATIONAL UNION OF PURE AND APPLIED CHEMISTRY (IUPAC)
and
INTERNATIONAL UNION OF BIOCHEMISTRY AND MOLECULAR BIOLOGY (IUBMB) Joint
Commission on Biochemical Nomenclature (JCBN)
Symbols for Specifying the Conformation of Polysaccharide Chains:
http://www.chem.qmul.ac.uk/iupac/misc/psac.html
5. Briefly about importance of carbohydrate-acting enzymes
- glycan binding proteins are abundant due to large amount of different carbohydrate structures
- for the synthesis or degradation of various glycan
molecules a large amount of specific enzymes are
needed
- specific for a type of bond and sugar molecules
connected by the bond
- wrong type of carbohydrate structures are oftentimes
found in cancer cells
29
6. Some extraordinary di- and oligosaccharides
(taken here just as examples of some practically important and interesting applications, but not
for exam):
- the hydroxy group in a sugar ring may also be derivatized
by another sugar ring, like in melezitose, which is a sugar
found as a minor component in honey
- sugar rings may also form ring-like structures
= cyclodextrins
- a-, b- and g-cyclodextrins have 6, 7 or 8 glucose units,
respectively
- cyclodextrins find use in applications like protecting
aroma molecules in foodstuffs or transferring and
delivering (slowly) drugs in the body due to complex
formation with such smaller molecules
- streptomycin, an antibiotic, interferes with bacterial protein synthesis
30
- bacterially produced dextrans
can be cross-linked to form very
hydrophilic preparations that
swell and form gels
- for example Sephadex, which is
used as a chromatographic
support media in gel filtration
(protein purifications)
31
- Sucralose is an artificial sweetener
Stevioside (steviol glycoside)
- natural sweetener extracted from
leaves of stevia plant
- 250-300 times sweeter than glucose
- both react with the taste receptors of the
tongue like sugars do, i.e. is sweet
- zero calories, because cannot be
metabolized in the body