Cellular Biology

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Cellular Biology
School of Life Sciences Shaanxi Normal University
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CHAPTER 5 EXTRACELLULAR MATRIX AND ORGANELLES
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Extracellular Matrix Extracellular matrix
(ECM) is a meshwork composed of macro molecules
secreted by cells
Collagen (Over 30)
Non-collagen glycoprotein
Aminoglycan and proteoglycan
Elastin
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Components of ECM
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The ECM of epithelial tissue
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Collagen The ratio of collagen is over 30
of total protein quantity in human body. Collagen
forms the meshwork in matrix. Collagen is
synthesized and transferred out by fibroblasts,
chondrocytes, osteoblasts, and some of epithelial
cells. So far, more than 20 types of
collagen have been established at least. The type
I, II, III, ?and ? are the fibro collagens that
have striation on them. The collagen I from
embryo or new born baby is easy to be extracted
because the conjugation between molecules is
inefficient. With aging, the conjugation is
increased and collagen I will become hard that
cause the hard skin, vascular tissue, that means
Old. Vitamin C is an important helper
factor to the synthesis of collagen I. The
inefficiency of vitamin C will cause blood
vessels easy to be broken and bleeding that we
call as Scurvy.
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Collagen fiber
Fibroblast
The collagen fibers surrounding fibroblast
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The types of collagen
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The structure of collagen L a model fig R an
image taken by electron microscope
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Fibronectin (FN) FN is a type of large
glycoprotein. FN can link cell on to ECM.
FN can exist in blood plasma or body
fluid(0.3mg/ml)as soluble type (Plasma FN) and in
ECM or cell surface as insoluble type (Cellular
FN). Plasma FN exists in a molecule formed a
dimer by two subunits linked at C terminal as a
V shape. Cell FN is a polymer. Some
short fragments of the FN chain are the smallest
structural units identified by FN receptors on
cell surface. For example, the RGD (Arg-Gly-Asp)
sequence existing in the cell binding domain
located in the middle of chain is the site that
can be recognized by some integrin receptors
located on cell surface.
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Model structure of FN
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FN links cell on to ECM
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Laminin (LN) LN is a large type of
glycoprotein too. Matrix is formed by LN and
collagen IV. LN is the earliest expressed
component of ECM during the embryonic
development. LN molecule is composed of one heavy
chain and two light chains as a cross shape.
At least, there are 8 cell binding sites in LN
molecule. For example, the 5 mers peptide, IKVAV
sequence, is located in the site near to the ball
domain of long arm that can bind to neuron cells
and enhance the neuron growth. 7 types of
LN molecules and 8 types of subunit
(a1,a2,a3,ß1,ß2, ß3,?1,?2) have been identified
so far. But, it is different from FN that each
subunit is encoded by different structural gene.
About 50 sugar chains are linked to the N
terminals of LN protein, so, LN is the most
complicated glycoprotein that we known so far.
Excepting LN and collagen IV, entactin,
perlecan, decorin and other proteins exist in
ECM.
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N terminals (Short arms)
C terminals (Long arm)
Model structure of LN
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The structure of ECM
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Glycosaminoglycan (GAG) GAG is a big family
of polysaccharides formed by repeated
disaccharide units. The features of GAG and
distribution GAG
Disaccharide unit Sulfate Distribution

radicals Hyaluronic acid
Glucuronic acid 0 Connective
tissue, skin
N-acetylglucose
Cartilage, vitreous body

Synovial fluid Chondroitin sulfate
Glucuronic acid 0.2-2.3 Cartilage,
bone, skin
N-acetylgalactose Cornea,
artery Dermatan sulfate Glucuronic acid
1.0-2.0 Skin, blood vessel
N-acetylglucose
Heart, cardiac valves Heparitin sulfate
Glucuronic acid 0.2-3.0 Lungs, artery,
cell surface
N-acetylglucose Heparin
Glucuronic acid 2.0-3.0 Lungs, liver,
skin,
N-acetylglucose
Mastocyte Keratin sulfate Galactose
0.9-1.8 Cartilage, cornea,
N-acetylglucose
Intervertebral discs
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L Proteoglycan M Polymers of proteoglycan R
Glycosaminoglycan
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Proteoglycan Proteoglycan is the compound
of GAG (except hyaluronic acid) and core protein
(Shown as the fig above). The polymer of
proteoglycan is composed of single proteoglycan
and hyaluronic acid. The molecule weight
of polymer of proteoglycan may be over 108KD, and
its volume may be bigger than bacteria.
Chondroitin sulfate and keratin sulfate are the
main members of GAG from the aggrecan that forms
cartilage. The inefficiency of the quantity of
the both GAGs can cause the inhibition of bones
development and short limbs and trunk. We call
these persons Dwarf. Elastin Elastin
meshwork makes tissue soft and elastic. The
extension of elastin is stronger 5 times than
rubber band at least. In the tissues of old
people, the synthesis of elastin is decreased and
the degeneration increased, that is why the
tissues of old people became hard, rough.
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Conjugation
Model structure of elastin
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• The functions of ECM
• Plays important role to the survival, growth, and
  death of cells. Eukaryotic cells must attach to
  ECM for their growth excepting blood cells, that
  we call as anchorage dependence. For example,
  epithelial cells will turn to apoptosis if they
  are separated from ECM. Different ECM will give
  cell different effects. The proliferation of
  fibroblast will be quick up on a fibronectin
  matrix, and slow down on a laminin matrix, but
  the response of epithelial cells to the matrix is
  just opposite.
• Controlls the shape of cell. Cells will be
  spherical if they grow with ECM separately. A
  cell can present a different shape if it grow on
  a different ECM. The mechanism of this regulation
  is mediated by the receptors on ECM that
  regulates the cytoskeleton.
• 3.Regulates the differentiation by the
  interaction between cell and special component of
  ECM. For example, myoblast (sarcoblast) can keep
  its original shape on fibronectin, but it will
  stop its proliferation to differentiate and fuse
  to myotube.
• 4.Mediates cell migration. ECM can regulate the
  speed and direction of the migration. Laminin can
  enhance the migration of tumor cells, and the
  migration of other cells is dependent on ECM too.
  This dependence is very important during the
  embryo development and wounds healing.
• So, ECM mediate almost every event in
  cell and life story.

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Cell proliferation
Cell survival
Cell growth
Cell migration
Cell shape
ECM
Apoptosis
Cell death
Cell differentiation
ECM plays role in almost each event of cell
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Organelles Many complicated
function areas are packaged by inner membrane
inside eukaryotic cells that we call as
organelle, or endomembrane system.
Organelles include nucleus, endoplasmic
reticulum, Golgi body (Golgi complex), lysosome,
mitochondrion, chloroplast, and others.
Every organelle is associated with some special
protein for protein synthesis, modification,
transportation or storage. Protein sorting
The proteins and lipids synthesized inside
cells must be transported into specific
organelles firstly, then transported out cells
like the follows
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Signal sequence or targeting sequence
Sorting receptor
Sorting receptor is the receptor located on
the endomembrane that can recognize the targeting
sequence of specific protein and transfer the
protein into organelle. Some organelle is the
place where proteins are modified, for example,
endoplasmic reticulum.
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Sorting signals for protein sorting Sorting
signals are the special peptide sequences or
pleated structures that can lead protein
directory transportation. Sorting signals lead
proteins transported into organelles from plasma,
or into plasma and ER from other organelles.
? Signal sequence A linear sequence existed in
the primary structure of protein that is composed
of 15-60 amino acids usually. Some of them will
be degenerated by signal peptidase after they
finished the directory transportation of protein.
Each signal sequence determines a specific
protein transportation direction. ? Signal
patch A pleated structure existed in the
tertiary structure of protein. The sorting signal
that leads protein into lysosome is signal patch.
Signal patch is difficult to be isolated from the
complicated tertiary structure of protein.
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Signal sequence and signal patch
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Some typical signal sequences
Functions Signal sequences
Into nucleus -Pro-Pro-Lys-Lys-Lys-Arg-Lys-Val-
Out of nucleus -Leu-Ala-Leu-Lys-Leu-Ala-Gly-Leu-Asp-Ile-
Into mitochondrion H3N-Met-Leu-Ser-Leu-Arg-Gln-Ser-Ile-Arg-Phe-Phe-Lys-Pro-Ala-Thr-Arg-Thr-Leu-Cys-Ser-Ser-Arg-Tyr-Leu-Leu-
Into plasma H3N-Met-Val-Ala-Met-Ala-Met-Ala-Ser-Leu-Gln-Ser-Ser-Met-Ser-Ser-Leu-Ser-Leu-Ser-Ser-Asn-Ser-Phe-Leu- Gly-Gln-Pro-Leu-Ser-Pro-Ile-Thr-Leu-Ser-Pro-Phe-Leu- Gln-Gly-
Into peroxisome -Ser-Lys-Leu-COO-
Into ER H3N-Met-Met-Ser-Phe-Val-Ser-Leu-Leu-Leu-Val-Gly-Ile-Leu-Phe-Trp-Ala-Thr-Glu-Ala-Glu-Gln-Leu-Thr-Lys-Cys- Glu-Val-Phe-Gln-
Back to ER -Lys-Asp-Glu-Leu-COO-(KDEL)
Into inside from plasma membrane Tyr-X-X-F
 
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• The ways by that proteins are sorted and
  transported
• Gated transportation
• For example, nucleopore can transport
  macromolecules selectively and automatically.
• Transmembrane transportation
• For example, passing through the
  translocator on the mitochondrion, the
  synthesized proteins located in plasma can be
  transported into mitochondrion under the leading
  of signal sequence with a linear molecule.
• Vesicular transportation
• Proteins are selectively packaged as
  vesicles and directively transported into target
  organelles. For examples, transportation from ER
  to Golgi body, formation of lysosome from Golgi
  body, adsorption of nutrition and hormone by
  cells.
• There are three capsid proteins can form
  vesicles to vesicular transportation clathrin,
  COPI and COPII.

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A Electron microscope images of clathrin
molecules B and C Models of capsid vesicle.
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Electron microscope images of clathrin capsid
vesicles
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Forming of clathrin capsid vesicle
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The direction of vesicles is dependent on
the marker protein and receptor of target. The
key proteins for that are SNAREs (soluble NSF
attachment protein receptor) and Rabs (targeting
GTPase). The fusion between HIV and CD4
is almost same to SNAREs way
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Adsorption of low density lipoproteins (LDL) by
cells LDL is very important to cause severe
cardiovascular diseases. Cholesterols can form
LDL (a vesicle actually) with phospholipids and
proteins. LDL are released into blood from liver
cells. The molecule weight of LDL vesicle is
3X106Da in 2030nm diameter. There are about 1500
cholesterol molecules packaged inside the
vesicle. So, the LDL outer layer is single layer
of lipid and a very huge protein (apolipoprotein
B-100) that can bind to the receptor of target
cell. Usually, cells will synthesize the LDL
receptors when they need cholesterol. But, if too
much cholesterol is accumulated in cell, cell
will stop the synthesis of receptor. As the
result, LDL level in blood will go up. For some
persons, a mutation of LDL receptor gene causes
their blood LDL level going up. LDL is much
easier to attach on to the inner wall of artery
than high density lipoprotein (HDL), especially
in brain and heart to form the atherosclerosis
that is number one killer to human being now in
the world.
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A model of LDL
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Endocytosis (vesicular transportation) of LDL
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Endoplasmic reticulum
Rough endoplasmic reticulum
(RER) Endoplasmic reticulum (ER)
Smooth
endoplasmic reticulum (SER) There are
ribosome particles attached on RER, and no any on
SER, that is why we see RER under microscope with
rough membrane and gave it the name, rough
endoplasmic reticulum. The function of ER
is to synthesize proteins and lipids.
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RER
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SER
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• Functions of ER
• Synthesis of proteins
• Usually, proteins synthesis is originally
  started in plasma and finished on ribosome. But,
  some proteins synthesis will be finished in RER
  after synthesis starting. For examples, ?
  secreted proteins, such as, antibodies, hormones
  ? transmembrane proteins ? Divided enzymes, such
  as hydrolases in lysosome ? Modified proteins,
  such as glycoprotein.
• There are 5 factors associated with the
  transferred synthesis at least
• ? Signal peptide A polypeptides that
  leads new synthesized peptide chain to ER, and it
  located at the N terminal of new synthesized
  peptide chain. Signal peptide is composed of
  1630 amino acids, and 6-15 of them are the
  positive charged and non-polarized amino acids.
  Signal peptide is named as start transfer
  sequence also.
• ? Signal recognition particle (SRP) SRP
  is composed of 6 different polypeptides, and
  binds to a 7S RNA. Its molecule weight is 325KD.
  SRP can bind to signal sequence to cause the
  protein synthesis paused.
• ? SRP receptor A integral protein can
  bind to SRP specifically.
• ? Stop transfer sequence A special
  peptide sequence with a high affinity to ER
  membrane. It can stop the peptide chain goes into
  ER and change the chain as a transmembrane
  protein.
• ? Translocator.

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Some sequences of signal sequences
Proteins Sequences of signal peptides
Preproalbumin Met-Lys-Trp-Val-Thr-Phe-Leu-Leu-Leu-Leu-Phe-Ile-Ser- Gly-Ser-Ala-Phe-Ser?Arg...
Pre-IgG light chain Met-Asp-Met-Arg-Ala-Pro-Ala-Gln-Ile-Phe-Gly-Phe-Leu- Leu-Leu-Leu-Phe-Pro-Gly- Thr-Arg-Cys?Asp...
Prelysozyme Met-Arg-Ser-Leu-Leu-Ile-Leu-Val-Leu-Cys-Phe-Leu- Pro-Leu-Ala-Ala-Leu-Gly?Lys...
2. Protein modification The modifications
include glycosylation (saccharification),
hydroxylation, acylation, and bisulfide bond
formation. The glycosylation is the most
important here. Glycosylation can ? have
proteins resistant to the digestion by enzymes of
digestion system ? give proteins markers,
signals, or cluster of determining (CD) ? have
some proteins pleated correctly. O-linked
glycosylation Link galactose or
N-acetylgalactosamine to the OH of Ser, Thr and
Hyp. The reaction is carried out on Golgi body.
N-linked glycosylation Link
N-acetylglucosamine to NH2 of Asn. The reaction
is carried out on ER.
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N-linked glycosylation
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3. Fold, assemble, and transportation of new
synthesized peptide chain. 4. Synthesis of
membrane lipids Most of membrane lipids are
synthesized in ER. Synthesized membrane lipids
will be transported to Golgi body and lysosome
with vesicular transportation way. 5.
Detoxication The P450 of SER is a monooxygenase
(mixed function oxidase or hydroxylase) that is
distributed on SER and other organelles. P450
can metabolize the fat soluble (liposoluble)
toxic substance into water soluble substance to
be ejected out of body. But, sometime, P450 can
activate the cancer inducer during the
detoxication. 6. Synthesis of steroid hormones
The enzymes in the SER, mitochondrion, Golgi body
of the endocrine cells of genital glands and
adrenal glands take duty to synthesize steroid
hormones. 7. Regulation to the level of blood
sugar. 8. Construction of some special
structures For example, the sarcoplasmic
reticulum specialized by the SER in muscle cells
can store Ca as the signal substance for the
muscle cell excitation.
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Golgi body
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The Golgi bodies in cultured epithelial
cells (Golgi body red Nucleus green)
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The enzymes in Golgi body include
glycosyltransferase, sulfo-glycosyltransferase,
redox enzymes, phosphatases, protein kinases,
mannosidase, transferases, and phospholipases.
Function regions Cis Golgi network (CGN) is
the entrance to receive the synthesized
substances and sort them into medial Golgi.
Medial Golgi is the place where the glycosyls
modification and glycolipids formation are
finished. Trans Golgi network (TGN) is the exit
to export the sorted and packaged proteins.
Each part of Golgi body has different feature
to cytochemical reactions ? Osmiophilic
reaction displays CGN. ? Cytochemical reaction
of thiamine pyrophosphatase (TPP enzyme)
displays TGN. ? Cytochemical reaction of
nicotinamide adenine dinucleotide phosphatase
(NADP or Co II enzyme) displays medial Golgi. ?
Cytochemical reaction of cytidylatase (CMP
enzyme) displays the vesicles and tubes closed
to TGN. CMP enzyme is the marker enzyme to
lysosome. Lysosome is manufactured in these
vesicles and tubes.
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CMP enzyme
The regions of Golgi body
TPP enzyme
NADP or Co II enzyme
Osmiophilic reaction
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Function of Golgi body 1. Glycosylation of
protein N-linked glycosylation is started on ER
and finished on Golgi body. Proteins will be
serially modified when they pass through the
Golgi body from Cis region to Trans region.
O-linked glycosylation is started and finished in
Golgi body. Glycosylation can mark protein,
change the structure of polypeptide, and
stabilize protein molecules. Proteoglycan is
formed in Golgi body also. 2. Exportation,
secretion and transportation of manufactured and
modified proteins Based on the signal peptide
and patch, Golgi body can sort synthesized
proteins on SER. Protein molecules CGN
Medial region (modified there) form vesicles
in TGN vesicles are fused with plasma of
TGN vesicles are released from TGN. 3.
Transformation of membrane New synthesized
membrane in ER can be transferred into Golgi body
to be remanufactured and modified, then,
transferred into plasma membrane with a
transporting vesicle that can be fused to plasma
membrane, that recruits plasma membrane. 4.
Hydrolyze proteins as active molecules The N or
C terminals can be cut off or hydrolyze protein
molecule as polypeptides. For examples, insulin
and neuropeptides. 5. Play role to form
lysosome. 6. Play role to form the cell wall of
plant. 7. Synthesize the cellulose and pectine
in cell wall of plant.
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Lysosome Primary
lysosome Lysosome Secondary lysosome
Residual body Primary lysosome
Primary lysosome is formed and secreted
by Golgi body. It obtains over 60 hydrolases
without activity. When the lysosome was broken or
substance else entered the lysosome, the enzymes
can be activated immediately. The hydrolases
include protease, nuclease, lipase, phosphatase,
sulfatase, phospholipases, and others. All of
them are the acidic hydrolases with a favorite pH
(5.0) to their activity. A proton pump
is located on lysosome membrane to input H into
cell for the low pH maintenance.
Lysosome membrane is highly glycosylated to
protect the membrane protein of themselves from
digested.
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Primary lysosome
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Secondary lysosome Secondary lysosome is
the lysosome that is digesting the substances
from outside of itself (phagolysosome) or inside
of itself (autophagolysosome).
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Residual body (post-lysosome) Residual body is
the lysosome that has lost its enzyme activity
and contains the undigested residues only.
Residual body can be exported out of cell, or
remained in cell, such as, lipofuscins in liver
cell.
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Function of lysosome Lysosome is a
digestion organelle in cell. It is associated
with cell autolysis, defense, and utilization to
some substances. Digestion in cell The
macromolecules entered by phagocytosis can be
digested by lysosome to be utilized by cell. For
example, the LDL can be digested for the sterol
utilization. Lysosome is unique digestion organ
in unicellular organism. Apoptosis The cell
that is going to apoptosis will form an apoptotic
body that can be swallowed by macrophage and form
a phagosome (secondary lysosome). The apoptotic
cell will be eliminated away from tissue by this
way. Autolysis To clear away the wasted bio
macromolecules and old organelles. Defense
Macrophage kills pathogens using its lysosome.
Regulation to endocrine Thyroglobulin can be
digested in lysosome as thyroxin. Formation
of acrosome of sperm Acrosome helps sperm to
enter ovum.
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Lysosome and diseases The damage,
function inhibition, non-digestion, gene mutation
or genetic deficiency of lysosome can cause many
diseases for human body, for examples, silicosis
(pneumosilicosis), pulmonary tuberculosis,
rheumatoid arthritis (The lysosome is easy to
break and release out the enzymes that can cause
the inflammation), and storage diseases (caused
by the mutation of the enzymes in lysosome). The
storage diseases include Tay-Sachs disease
(?-?????), Pompe disease (II??????), Gaucher
disease (??????), Inclusion-cell disease
(??????), and others. All patient children will
die within two years. Peroxisome (microbody)
0.21.5µm in diameter. Over 40 oxidases
have be found in peroxisome. Oxidases can oxidize
substrates and release out hydrogen peroxide
(H2O2)
RH2O2 ? RH2O2 Peroxisome take
the ß-oxidation of the fatty acids in animals. If
you use some medicine of fat cleaner to rat, you
will detect a 10 folds higher concentration of
oxidases in rat liver cells than normal.
Peroxisome can take a detoxifying function that
oxidizes the toxic substances with H2O2. For
example, about a quarter of the alcohol you drank
in will be oxidized as acetaldehyde in your liver
cells. Plant peroxisome can ? join
photorespiration ? take ß-oxidation of fat.
The signal to lead oxidases from plasma
enter peroxisome is -Ser-Lys-Leu-COO-?
The syndrome of Zellweger (??????) is caused by
abnormal peroxisome (blank peroxisome). The
infant will die within 3-6 months.
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Peroxisomes in human liver cells
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The peroxisome in the cell of Nicotiana tabacum
(The central square is the crystal formed by
uricoxidase)