q l w ¦ ¥ w § cr k d ²seika/h24/_userdata/...é í ç è ï ç ê ï ñ question 16-2 essential...
TRANSCRIPT
Essential
Molecular Biology of the Cell 6th editionGarland Publishing, Inc.
A
etc.
M78Figure 11-3 Essential Cell Biology (© Garland Science 2010)
50 µm : 5 nm = 10 m : X
X = 1 mm
2-
2- 2-ME
Urea
HS-CH2-CH2-OH
O=CCH3
CH3
--
NH2+=C
CH3
CH3
--
α β
NH2I1IMSSVQSQQEQLSQSDPSPSPNSCSSFELIDMDAGSLYEPVSPHWFYCKIIDSKETWIPFNSEDSQQLEEAYSSGKGCNGRVVPTDGGRYDVHLGERMRYAVYWDELASEVRRCTWFYKGDKDNKYVPYSESSSQVLEETYMLAVTLDEWKKKLESPNREIIILHNPKLMVHYQPVAGSDGWGSTPTEQGRPRTVKRGVENISVDIHCGEPLQIDHLVFVVHGIGPACDLRFRSIVQCVNDFRSVSLNLLQTHFKKAQENQQIGRVEFLPVNWHSPLHSTGVDVDLQRITLPSINRLRHFTNDTILDVFFYNSPTYCQTIVDTVASEMNRIYTLFLQRNPDFKGGVSIAGHSLGSLILFDILTNQKDSLGDIDSEKDSLNIVMYQGDTPTLEEDLKKLQLSEFFDIFEKEKVDKEALALCTDRDLQEIGIPLGPRKKILNYFSTRKNSMGIKRPAPQPASGANIPKESEFCSSSNTRNGDYLDVGIGQVSVKYPRLIYKPEIFFAFGSPIGMFLTVRGLKRIDPNYRFPTCKGFFNIYHPFDPVAYRIEPMVVPGVEFEPMLIPHHKGRKRMHLELREGLTRMSMDLKNNLLGSLRMAWKSFTRAPYPALQASETPEETEAEPESTSEKPSDVNTEETSVAVKEEVLPINVGMLNGGQRIDYVLQEKPIESFNEYLFALQSHLCYWESEDTVLLVLKEIYQTQGIFLDQPLQI711ICOOH
Ap5A
5
Aβ
PrP
PrP
PrPsc
500
75
PrP
PrP
PrP
PrP
PrPsc PrPc PrPsc
PrPsc β
1997
Figure 4-8 Essential Cell Biology (© Garland Science 2010)
PrpC Prpsc
Prpsc
PrpC PrpSC
PrpC PrpSC
α- α- β-
APP
APP
Figure 10-20 Molecular Biology of the Cell (© Garland Science 2008)
#
DNA
DNA → mRNA →
DNA → mRNA →
2013
autocrine
YY
Question 16-2 Essential Cell Biology (© Garland Science 2010)
Histamine
H1-H4 GPCR
マスト(肥満)細胞�
・ 蕁麻疹・ 喘息・ 花粉症・ アトピー性皮膚炎�
・ ヒスタミン・ LTC4, LTD4・ TXA2・ PGD2・ プロテアーゼ・ IL-5・ TNF-α
メディエーター�
関連病態�
・ 粘膜�・ 皮膚・ 腸管�・ 血管周囲�
存在部位�
抗原
ヒスタミン Leukotriene PAF ThromboxaneIL-5 TNF-α プロテアーゼ
マスト細胞
IgE 受容体IgE
マスト細胞は IgE-抗原に反応し、脱顆粒する
Figure 25-1 Molecular Biology of the Cell (© Garland Science 2008)
Figure 25-2 Molecular Biology of the Cell (© Garland Science 2008)
T
Figure 25-3 Molecular Biology of the Cell (© Garland Science 2008)
Figure 25-6 Molecular Biology of the Cell (© Garland Science 2008) Figure 25-7a Molecular Biology of the Cell (© Garland Science 2008)
Figure 25-7b Molecular Biology of the Cell (© Garland Science 2008) Figure 25-7c Molecular Biology of the Cell (© Garland Science 2008)
Figure 25-2 Molecular Biology of the Cell (© Garland Science 2008)
T
Figure 25-19 Molecular Biology of the Cell (© Garland Science 2008)
Table 25-1 Molecular Biology of the Cell (© Garland Science 2008) Figure 25-21 Molecular Biology of the Cell (© Garland Science 2008)
Figure 25-23 Molecular Biology of the Cell (© Garland Science 2008)
144
THE ANTIBODY MOLECULE B CELLS PRODUCE ANTIBODIES
ANTIBODY SPECIFICITY
Antibodies are proteinsthat bind very tightly totheir targets (antigens).They are produced invertebrates as a defenseagainst infection. Eachantibody molecule ismade of two identicallight chains and twoidentical heavy chains,so the two antigen-binding sites areidentical.
Antibodies are made by a class of white blood cells called Blymphocytes, or B cells. Each resting B cell carries a differentmembrane-bound antibody molecule on its surface that servesas a receptor for recognizing a specific antigen. When antigenbinds to this receptor, the B cell is stimulated to divide and tosecrete large amounts of the same antibody in a soluble form.
RAISING ANTIBODIES IN ANIMALSAntibodies can be made in the laboratory by injecting an animal(usually a mouse, rabbit, sheep, or goat) with antigen A.
Repeated injections of the same antigen at intervals of several weeks stimulate specific B cells to secrete large amounts of anti-A antibodies into the bloodstream.
Because many different B cells are stimulated by antigen A, theblood will contain a variety of anti-A antibodies, each of whichbinds A in a slightly different way.
antigen-binding sites
light chain
heavy chainhinge
5 nm
antigen
heavy chain
light chain
An individual animalcan make billions ofdifferent antibodymolecules, each with adistinct antigen-bindingsite. Each antibodyrecognizes its antigenwith great specificity.
different B cells
Antigen binds toB cell displaying anantibody that fitsthe antigen.
B cell is stimulated to make and secrete more of same antibody.
inject antigen A take blood later
A
A A Atime
amou
nt o
f an
ti-A
anti
bodi
es in
blo
od
ANTIBODIES DEFEND US AGAINST INFECTION
foreignmolecules
viruses bacteria
ANTIBODIES ( ) FORM AGGREGATES
Antibody and antigenaggregates are ingested
by phagocytic cells.
Special proteins inblood kill antibody-
coated bacteriaor viruses.
PANEL 4–3 Making and using antibodies144
THE ANTIBODY MOLECULE B CELLS PRODUCE ANTIBODIES
ANTIBODY SPECIFICITY
Antibodies are proteinsthat bind very tightly totheir targets (antigens).They are produced invertebrates as a defenseagainst infection. Eachantibody molecule ismade of two identicallight chains and twoidentical heavy chains,so the two antigen-binding sites areidentical.
Antibodies are made by a class of white blood cells called Blymphocytes, or B cells. Each resting B cell carries a differentmembrane-bound antibody molecule on its surface that servesas a receptor for recognizing a specific antigen. When antigenbinds to this receptor, the B cell is stimulated to divide and tosecrete large amounts of the same antibody in a soluble form.
RAISING ANTIBODIES IN ANIMALSAntibodies can be made in the laboratory by injecting an animal(usually a mouse, rabbit, sheep, or goat) with antigen A.
Repeated injections of the same antigen at intervals of several weeks stimulate specific B cells to secrete large amounts of anti-A antibodies into the bloodstream.
Because many different B cells are stimulated by antigen A, theblood will contain a variety of anti-A antibodies, each of whichbinds A in a slightly different way.
antigen-binding sites
light chain
heavy chainhinge
5 nm
antigen
heavy chain
light chain
An individual animalcan make billions ofdifferent antibodymolecules, each with adistinct antigen-bindingsite. Each antibodyrecognizes its antigenwith great specificity.
different B cells
Antigen binds toB cell displaying anantibody that fitsthe antigen.
B cell is stimulated to make and secrete more of same antibody.
inject antigen A take blood later
A
A A Atime
amou
nt o
f an
ti-A
anti
bodi
es in
blo
od
ANTIBODIES DEFEND US AGAINST INFECTION
foreignmolecules
viruses bacteria
ANTIBODIES ( ) FORM AGGREGATES
Antibody and antigenaggregates are ingested
by phagocytic cells.
Special proteins inblood kill antibody-
coated bacteriaor viruses.
PANEL 4–3 Making and using antibodies
IgM
IgG
IgM
Figure 25-25 Molecular Biology of the Cell (© Garland Science 2008) Figure 25-26 Molecular Biology of the Cell (© Garland Science 2008)
抗原
ヒスタミン Leukotriene PAF ThromboxaneIL-5 TNF-α プロテアーゼ
マスト細胞
IgE 受容体IgE
マスト細胞は IgE-抗原に反応し、脱顆粒する
ADCC (Antibody-Dependent-Cellular-Cytotoxicity CDC Complement-Dependent Cytotoxicity :
•
•
•
PD-1, PD-L1
Figure 25-43 Molecular Biology of the Cell (© Garland Science 2008)
B
Y Y
Y
Y
Y Y
Y
Y Y
Y
Y
B
Y Y
Y
Y
Y YY Y
Y
YY Y
Y
Y
Y YY Y
Y
Y
Y Y
Y
Y
Y YY Y
Y
Y
Y Y
Y
Y
Y YY Y
Y
Y
Y Y
Y
Y
Y YY Y
Y
Y
Y Y
Y
Y
Y YY Y
Y
Y
145
USING ANTIBODIES TO PURIFY MOLECULES
MONOCLONAL ANTIBODIES USING ANTIBODIES AS MOLECULAR TAGS
M A A
A
A A A A
A
E P F N O R D
S C
J L B K
Q G H
MA
AA
E P
F
NOR D
SCJ
J
L
B
B
KQG H
IMMUNOPRECIPITATION IMMUNOAFFINITY COLUMN CHROMATOGRAPHY
mixture of molecules
add specific anti-A antibodies
collect aggregate of A molecules andanti-A antibodies by centrifugation
mixture of molecules
discard flow-through wash collect pure antigen A
washelute
antigen A
bead coated with anti-A antibodies
column packed with these beads
Large quantities of a single type of antibodymolecule can be obtained by fusing a B cell(taken from an animal injected with antigen A)with a tumor cell. The resulting hybrid cell divides indefinitely and secretes anti-A antibodies of a single (monoclonal) type.
Tumor cell fromcell culture dividesindefinitely butdoes not makeantibody.
B cell from animalinjected with antigenA makes anti-Aantibody but doesnot divide forever.
FUSE ANTIBODY-SECRETINGB CELL WITH TUMOR CELL
Hybrid cellmakes anti-Aantibody anddividesindefinitely.
couple to fluorescent dye,colloidal gold particle, orother special tag
specific antibodiesagainst antigen A labeled antibodies
Fluorescent antibody binds toantigen A in tissue and isdetected by fluorescence in a light microscope. The antigen here ispectin in the cell walls of a sliceof plant tissue.
Gold-labeled antibody bindsto antigen A in tissue and is detected in an electron micro-scope. The antigen is pectinin the cell wall of a singleplant cell.
Antigen A isseparated fromother moleculesby electrophoresis.
Incubation with thelabeled antibodiesthat bind to antigen Aallows the position of theantigen to be determined.
Note: In all cases, the sensitivity canbe greatly increased by using multiplelayers of antibodies. This “sandwich” method enables smaller numbers of antigen molecules to be detected.
antigen
Labeled second antibody (blue) binds to first antibody (black).
C KNR
A
A
A
A
BA
A
MIC
ROSC
OPI
C D
ETEC
TIO
NBI
OCH
EMIC
AL
DET
ECTI
ON
etc
J
cellwall
Fab
抗体フォーマットの変換
Bivalent Mini-Antibody(Fab_dHLX_MH)
SDS-PAGESDS
Western blot
DetergentSDSTriton X-100Nonidet P-40
Triton X-100
Tween20
n-
Sodium Dodecyl Sulfate
SDS-PAGE
50
37
kDa
NPP6
NPP6
MBP
*
*
**
**
* ** Coronal section Magnified view
0.2 mm 5 µm 5 µm
X NMR
Chapter 10 Analyzing Genes and Genomes
Copyright © Garland Science 2010 Copyright © Garland Science 2010
Chapter 10 Analyzing Genes and Genomes
Copyright © Garland Science 2010
GPCR
X
A
A
PCR
Cytomegalo virus SV40
A
PCR: Polymerase Chain Reaction
RT-PCR: Reverse Transcriptase PCR
mRNAcDNA
mRNA A
× ×
RNA
PCR RT-PCR cDNA
cDNA
RNA
PCR RT-PCR cDNA
cDNA
X
GFP GFP
GFP
mcherry
DNA分子の変性と再生を利用した手法
mRNA
DNA
in situmRNA DNA
#CRISPR TALEN
,
GPCR
T