The Elaborate Structure of Spider Silk

Report
The Elaborate Structure of
Spider Silk
Grace Hii Leh Ung
Ng Si Ling
Yeow See Leong
Lee Wen Hau
Tan Teng Teng (Jennise)
Fasihah binti Che Muni
Khairun Nazihah binti Khalid
B050810211
B050810253
B050810105
B050810183
B050810016
B050810093
B050810081
1
Introduction
Outstanding
mechanical
properties
Antimicrobial &
hypoallergenic
Spider
Silk
Fiber
Eco-friendly
& sustainable
nature
Unique self
assembly
2
Introduction (cont.)
 Strong interest in advanced composite industry.
 Ultra-lightweight fiber that combine enormous
tensile strength with elasticity.
 Each fiber stretch up to 40% of its length and
absorb hundred times as much energy as steel
without breaking.
 High antimicrobial and hypoallergenic properties
lead to low infection rate from inflammation and
allergic reaction in application of biomaterials.
3
Different silk types produced by female orb weaving spiders (Araneae). Each silk
type (highlighted in red) is tailored for a specific purpose.
Eisoldt,L., Smith,A. and Scheibel,T., 2011. Decoding the secrets of spider silk. Materials Today, Vol. 14, no. 3, pp.8086.
4
Drag-line Silk
 Used by spider for frames of their webs and as safety
lines.
Structures:
 Made up of crystalline regions of anti-parallel ß-sheets
and “non-structured amorphous regions (coiled coil,
performed ß-sheets and elastic ß-turn spirals).
 Crystalline arrays responsible for stiffness of fiber.
 Amorphous regions (55 to 60% of dragline spider silk)
more-or-less kinetically free and change shape under
influence of external load and through entropic elasticity
5
Properties:
 Stronger than high tensile strength steel and approaches
stiffness and strength of high performance p-aramid fiber
KEVLAR (widely used in bulletproof vests).
 Higher toughness due to greater extensibility.
 Torsional dampening behavior. When dragline thread
twisted, it does not oscillate around the new position,
like a Kevlar fiber would. After a while, fiber slowly
returns to its initial position
shape memory within
fiber.
 Ability to undergo supercontraction. When dragline silk is
wetted, or when the relative humidity is above 60 %, a
silk thread swells in diameter and shrinks in length by
about 50 %.
6
Primary Structure of Spider Silk
• Primarily consists of protein that possess large
quantities of non-polar and hydrophobic
amino acids (glycine or alanine).
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Primary Structure of Spider Silk (cont.)
• Furthermore, it contain highly repetitive amino acid sequence
(>90%), especially in their large core domain.
• Composed of short polypeptide stretches of about 10-50
amino acids.
• Can be repeated more than a hundred times within one
individual protein.
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Primary Structure of Spider Silk (cont.)
• MA and Flag silks contain up to four typical aligopeptide motifs:[I]
(GA)n/(A)n, [II] GPGGX/GPGQQ, [III] GGX (X = A, S or Y) and [IV] “spacer”
9
Primary Structure of Spider Silk (cont.)
• Nonrepetitive regions are located at the protein’s termini (comprise
approx. 100-200 amino acids)
• The N-terminus (NRN –domain) refers to the start of
a protein or polypeptide terminated by an amino acid with a
free amine group (-NH2).
• The C-terminus (NRc –domain) is the end of an amino acid chain
(protein or polypeptide), terminated by a free carboxyl group (-COOH).
• Due to cysteine residues, Intermolecular disulfide bonds stabilize these
tertiary structures of proteins.
• These domains are thought to initiate and specify assembly of silk
proteins.
10
Primary Structure of Spider Silk (cont.)
• The primary structure shows a specific hydrophobicity pattern with
alternating hydrophilic and hydrophobic blocks in their core domain.
• Such amphiphilic (have both hydrophobic and hydrophilic domains)
composition is reminiscent of surfacetant or biological membranes.
• For the case of spider silks, it is important for phase separation during the
spinning process.
• Amphiphilic pattern might be responsible for formation of micelles
postulated as intermediate structures during thread assembly.
A droplet of water forms a spherical shape to
minimize contact with the hydrophobic leaf.
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Model of the Silk Spinning Process
12
• Highly Concentrated (Monomers ) Spider silk protein solution is secreted
Spinning
and stored inside the spinning gland.
gland
• Phase separation take place when aligning in one direction through a
Spinning
narrow ion exchange channel- formation liquid crystalline behavior of the
Duct
spinning dope.
• Loss of solvent the conformation conversion is finalized –silk fiber is
Spinning
drawn out to form silk thread.
wart
13
• Assemble into small micelles with a diameter ~ 100-200nm due to
amphiphilic properties inside the spinning dope.
Spinning
• Multimerization occurs forms a globules in micrometer range.
gland
Spinning • Through laminar force , the shear force increase due to narrow channel.
Duct
Spinning • Force these globules into an elongated shape-formation of fiber (silk thread).
wart
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Mechanical Properties
• The most outstanding property of spider silk
 maximal resilience
• Able to absorb energy 3X more than Kevlar
• Kevlar is one of the sturdiest material.
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• Resilience  the ability of a material to absorb
energy when it is deformed elastically and release
that energy upon unloading.
• Resilience = area under the stress-strain graph
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• Spider silk : well-balanced between strength &
elasticity
17
• Synthetic fibers  higher stiffness & strength than natural fibers
• Natural fibers tend to be more elastic
• Synthetic carbon fibers: yield point > 5X higher than the best
insect silk.
18
Mechanical properties of spider silk from Nephila clavipes
compared to other structural materials.
Material
Strength (Nm-2)
Elasticity (%)
Energy to break (J
kg-1)
Dragline silk
4 x 109
35
1 x 105
Flagelliform silk
1 x 109
>200
1 x 105
Kevlar
4 x 109
5
3 x 104
Rubber
1 x 106
600
8 x 104
Tendon
1x 109
5
5 x 103
Nylon, Type 6
4 x 107
200
6 x 104
Fischer,R. and Schillberg,S., 2006. Molecular Farming: Plant-Made Pharmaceuticals and Technical Proteins.
Germany: John Wiley & Sons, p. 172.
19
• Spider silk shows high supercontraction!!
Absorption of water  50% shrinkage  and
behaving like a weak rubber under tension 
tightens the thread  ensure rigidity
Stronger, stiffer, less extensible and better able
to recover after being stretched
The presence of MaSp2 rich in GPGXX
motifs increases the capacity of MA silk to
supercontract
Water disrupts the hydrogen bonds that hold
the GPGXX motifs and 310 helices within the
silk parallel
the GPGXX motifs and 310 helices rearrange
to a lower energy state and the fiber loses its
orientation
20
Highest Performance Spider Silk
• The toughest known spider silk is produced by
the species Darwin’s bark spider (Caerostris
darwini)
• Average toughness = 350 MJ/m3 with some
samples reaching 520 MJ/m3.
• C. darwini silk is more than twice as tough as
any previously described silk and over 10
times tougher than Kevlar
21
Darwin’s bark spider (Caerostris darwini)
22
Mimicking Nature
 Define as evolve to share common
perceived characteristics with another group,
the models. The evolution is driven by
the selective action of a signal-receiver, or dupe.
 The signal-receiver is typically another
intermediate
organism
like
the
common predator of two species, but may
actually be the model itself, such as a moth
resembling its spider predator.
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1. Recombinant spider silk
Method
 Direct transformation of original or fragmented
silk genes into bacterial hosts.
 Depends on gene encoding a protein from one
organism being transferred into a production host
such as the bacterium Escherichia(E.) coli.
 Spidroin genes have an extreme codon bias, with
only a subset of available codons being used.
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1. Recombinant spider silk (cont.)
 Issues
1. Homologue recombination in many hosts, which removes
repetitive sequences which resulted in extremely low yields
of the intended protein.
2. Direct transformation of original silk genes and silk
fragments is not the method of choice for recombinant
spider silk production.
 Solutions
1. genes coding were generated using a cloning strategy which
is based on a combination of synthetic approach.
2. Over-expression of the gene in bacterial hosts encoding DNA
which increased DNA level, it is possible to achieve much
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higher silk gene in E. coli.
2. Artificial spinning of spider silk
 Made of hundreds of tubes coming from silk
glands called major ampullate and minor
ampullate, and the number of glands of spider
varies with species.
 Due to availability of recombinant spider silk
proteins, scientists able to analyze assembly of
spider silk threads in a functional in vitro spinning
process in the near future which ensure the
generated silk fiber resembles natural silk in its
microstrucuture, chemical, composition and
mechanical properties.
26
Method
1. Wet-spinning processes employed with promising results.
Using silicon micro-spinnerets, several meters of insect or
spider silk fibers produced.
Natural and artificial spinning ducts.
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2. Artificial spinning of spider silk
(cont.)
2. Special Postspinning techniques yielded silks
even larger diameters.
But, the best mechanical properties obtained by
artificial spinning techniques are much lower
than natural dragline silks.
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… Q & A …
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1. What are the applications of spider silk?
• Military: bulletproof vest
 Polymer science. Eg: polycarbonate (PC)-improve with
adding of spider silk.
 High toughness
 Able to dissipate energy at high strain rates
Ultimate tensile
strength
Elongation
Spider
silk
PC
11 GPa
72 GPa
27%
6%
30
• Medical: wound healing
Minimal risk for infection or disease
transmission
Biocompatibility for implanted
material (implantation)
Immediate ligament stabilization
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• Construction: Scaffolding
 Lightweight
 High toughness
 High strength
 Can stand with temperature in below -40°C up
to 220°C
32
2. Why goat is chosen in producing spider silk?
• Not many differences between the silk glands of
spiders and the milk glands of goats, it should be very
easy to produce the silk in goats milk.
33
3. How to produce spider-goat silk?
• The silk-making genes of a spider and the genome of
goats is spliced.
• The genes were altered to only turn on in the
mammary glands of female goats that are lactating.
• Using cloning techniques they have bread goats that
have the spider genes, produce female goats that
have the silk gene and can produce the silk in their
milk.
• When the milk is produced the silk proteins will be
isolated out of the milk, and purified, this produces a
whitish liquid that can then be spun into fibers and
used for the many applications.
34
35
References
Fischer,R. and Schillberg,S., 2006. Molecular
Farming: Plant-Made Pharmaceuticals and
Technical Proteins. Germany: John Wiley & Sons,
p. 172.
Eisoldt,L., Smith,A. and Scheibel,T., 2011.
Decoding the secrets of spider silk. Materials
Today, Vol. 14, no. 3, pp.80-86.
Ciement,G., 2011. Fundamentals of Space
Medicine, Second Edition. London: Springer, p.69.
36
J. M. Gosline, P. A. Guerette, C. S. Ortlepp And
K. N. Savage. (1999). The Mechanical Design
Of Spider Silks: From Fibroin Sequence To
Mechanical Function, p. 3295-3303
 Cecilia Boutry and Todd Alan Blackledge.
(2010). Evolution of Supercontraction In
Spider Silk: Structure–Function Relationship
from Tarantulas to Orb-Weavers, p. 3505-3514
 Mark J. Bonino. (2003). Material Properties of
Spider Silk, p. 21-54
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