林光輝 :

生化科教授

醫學大樓 Room 0861Tel: 5177, 5975

e-mail:khlin@mail.cgu.edu.tw

上課投影片網址

長庚大學—系所介紹—基礎醫學研究所 -生物化學暨細胞分子生物學組 -師資介紹-林光輝-講義上網-Biochem

C Chapter 8 Nucleotides and Nucleic Acids  Basic information: Griffith--pneumococcus bacteria; Avery,MacLeod, MacCarty (1944)* Hershey, Chase - bacteriophages (1952)* Chargaff - ratios of purines to pyrimidines = 1,

A/T=1, G/C=1 (late 1940s) Wilkins, Franklin - X-ray diffraction provided clues to

the shape of the DNA molecule (1950)* Watson, Crick-double helix (1953)

X-ray diffraction :Franklin and Wilkins

Nucleic acids: monophosphates: Base + sugar + phosphate  H-bond propertiesUV-light absorbing properties Sugars: 5-Carbon pentose: D-ribose: RNA 2'-deoxy-D-ribose: DNA Nucleoside: N-glycosidic linkage of base to carbon of sugar

Nomenclature Pyrimidines:+ idine: Cytidine, uridine,

thymidine Purines: + osine: Adenosine, guanosine  Nucleotides: Typically 5' nucleoside phosphate

5‘-ribonucleoside monophosphates AMP, GMP, CMP, UMP: cAMP, cGMP: 3',5'-Cyclic ribonucleoside monophosphates: Regulation of cellular metabolism

5'-Ribonucleoside diphosphates

ADP, GDP, CDP, UDP:

5’-Ribonucleoside triphosphates

ATP (energy currency),

GTP (protein synthesis and signal transduction),

CTP (lipid synthesis),

UTP (carbohydrate and polysaccharide synthesis)

5’-Deoxyribonucleoside- triphosphates

dATP, dGTP, dCTP, dTTP: For DNA synthesis

Structure of nucleotides

N-glycosidic bond

Nomenclature

Base Nucleoside Abbreviation

adenine adenosine Aguanine guanosine Gcytosine cytidine Curacil uridine Uthymine thymidine T

Important to know:

U in RNA onlyT in DNA only

More Nomenclature

Nucleoside = base + sugar

Nucleotide = base + sugar + phosphate

5-methycytosine is a mutation hot spot

Double stranded DNA is written with the 5’ to 3’ on top

5’ - GATC - 3’3’ - CTAG - 5’

GATCCTAG

=

If only one strand of DNA is given, it is assumed to be in the 5’ to 3’ orientation

GATC = GATCCTAG

DNA replication

Double stranded DNA: Strands antiparallel:

Base pairs AT and GC

*B-form DNA: Right-handed helix

Parameters:0.34 nm/base pair, 10 bp/turn

H-bonded base pairs AT and GC inside

Base pairs perpendicular and stacked

Negative charged phosphate and sugar outside

Major and minor groove

*A-form DNA: Right-handed helix

Parameter: 0.224 nm/base pair, 11 bp/turn

Base pairs tilted

dsRNA and DNA/RNA hybrids

*Z-form DNA: Left-handed helix

Dinucleotide repeat: Pyrimidine-purine

Conformation: Purine -syn, pyrimidine -anti (p284)

Zigzag pattern of sugar-phosphate backbone

Gene regulation or in genetic recombination

Adenosine: A mediator of the sleeping inducing effect of prolonged wakefulness

Function:1. Blood vessel dilation.2. Slow the heart rate (adenocardTM)具有強力的抗心律不整及冠狀動脈擴張作用。

臨床上則是利用其抗心律不整之作用。3. Sleepiness

Peptide nucleotide (PNA)

Sugar phoaphate backbond—NAPeptide backbond—PNA

Function:1. Resistant to nuclease or protease2. Diagnostic probes3. Antisense drugs

Nucleic acids: Nucleoside monophosphates in phosphodiester linkage 5' to 3'

 DNA: Genetic material: double stranded

dsDNA: Strands antiparallel

Interchain H bonds form base pairs

Chargaff's rules: A=T, G=C, Purines = Pyrimidines

X-ray diffraction of Franklin and Wilkins and model building of Watson and Crick

1/1

What is RNA?

Ribonucleic acid– Ribonucleotides (Ribose, base, & phosphate)

Types– Coding: messenger RNA (mRNA)– Non-coding:

• Ribosomal RNA (rRNA)• Transfer RNA (tRNA)• Small nuclear RNA (snRNA)• Small nucleolar RNA (snoRNA)• Interference RNA (RNAi)• Short interfering RNA (siRNA)• Micro RNA (miRNA)

MCB 1.6

RNA can form secondary structuresby internal base-pairing

RNA can also be used:

- to store genetic information (viruses)

-for catalytic activity (self-splicing, RNaseP)

-Regulation

RNA:Typically single stranded: Produced during transcription

mRNA: Carries information encoded in genes to direct protein synthesis on ribosomes

*Derive from heterogeneous nuclear RNA (hnRNA)

mature mRNA processed by splicing (removal of introns and joining of exons), capping (5'

end), and polyA tail addition (3' end)

MCB 3.15

Basic differences between Eucaryotes and Procaryotes

(Bacteria)

DNA Pol vs RNA Pol

DNA replication Gene transcription Ori promoter dNTP NTP Proofreading NO Replicate dsDNA Only template Primer require NO

Different at transcriptional level ofProkaryotic vs Eukaryotic cell

One RNA Pol Three Continuous gene Discontinuous gene NO intron/Extron Yes mRNA unstable more stable No processing Yes(capping,polyA addition,splicing) Trancription/Translation NO coupled Polycistron Moncistron

rRNA Components of ribosome: Protein synthesis

Small subunit: Single rRNA: 16S (1500 nt) or 18S (1800 nt)

Large subunit: Two or three

23S (2900 nt) or 28S (5000 nt),

5S, 5S rRNA: 120 nt,

5.8S rRNA: 160 nt (eukaryotic cytoplasmic ribosomes).

Blue color: Prokaryotic; Pink, eukaryotic cell.

tRNA:carriers of activated amino acids used by ribosome for protein synthesis.

*Small (73 to 94 nucleotides) RNA charged with amino acids

* Cloverleaf secondary structure

*Acceptor stem, CCA 3’: end at which amino acid is attached

*Anticodon stem and loop: three-base anticodon: decodes mRNA

tRNA

DNA vs RNA

Composition:

* T in DNA not U to distinguish from T formed by deamination of C

* 2' OH in RNA accounts for instability of RNA

 

phosphodiester bond Hydrolysis

*RNA:Sensitive to base hydrolysis but resistant to acid hydrolysis

*DNA:Resistant to base hydrolysis: Depurinates by acid hydrolysis (apurinic base)

DNA denaturation: Strand separation and loss of

base stacking

Temperature, pH, ionic strength: Disrupt H bonds

Hyperchromic shift of UV absorbance upon denaturation

Tm: Midpoint of transition.

 

Renaturation

* Second order kinetics, * Nucleic acid hybridization

Measure evolutionary relationships

Identify specific gene

  

2/1C

Affect by:

PHIonic strengthSizeGC content

Stability of NA

dsRNA>RNA/DNA>dsDNA

PH7.0 denaturation of dsRNA is about 200C higher than denaturation of dsDNA

Rotator; Nurses run

Replication, recombinationTranscription.

The N-7, O-6, and N-6 of purines base-paringForm triplex DNAs, stable at low PH, C=G.C+

Hoogsteen pairing:

Quadruplex: DNA sequences with very high G

H-DNA: polypyrimindine or polypurineRegions involved in gene regulation.

Deamination

Allopurinol: Xanthine Oxidase inhibitor

Depurination: DNA, pH 3, purine base removed

A potent reagent of theDeamination of bases.Food preservativesDNA damage

Alter baes of DNAMethylation of G

2/1M

DNA sequencing

*Chain termination (dideoxynucleotide): Sanger

Enzymatic synthesis of DNA using DNA polymerase

Primer extended using dNTPs but terminated using specific ddNTPs (lack 3' hydroxyl)

Four reactions produce four sets of products

Analyze products by electrophoresis

* Automatic DNA sequencing: End-label DNA with dideoxynucleotides with fluorescent dyes of different colors

Adenine nucleotides are components

of many enzyme cofactors:

Adenine nucleotide are components of many enzyme cofactors

single nucleotide polymorphism:

DNA single base variations found in more than 1% of population.

SNP

What is a SNP?What is a SNP?What is a SNP?What is a SNP?

Genetic DiversityGenetic DiversityGenetic DiversityGenetic Diversity

0.3%

10 million SNPs

Human Variations

0.3%

2%

SNP: single nucleotide polymorphism

•DNA single base variations found in >1% of population.

•Abundant: ~3 million per individual genome.

•Distribution throughout the genome and with high density.

• Most common form of genetic variation-Genetic linkage studies-Genome-wide association studies

• Indicate predisposition to-Disease predisposition and onset-Drug tolerance-Drug efficacy

• Genome SNP scans will uncover gene function, and define new drug targets

• SNPs will enable physicians to personalize therapy

Why are SNPs Important?Why are SNPs Important?Why are SNPs Important?Why are SNPs Important?

1. Even SNPs that do not change protein expression and cause disease may be close on the chromosome to deleterious mutations.

2. Because of this proximity, SNPs may be shared among groups of people with harmful but unknown mutations and serve as markers for them.

SNP

SNP

Analyzing shifts in SNPs among different groups of people will help population geneticists to trace the evolution of the human race down through the millennia and to unravel the connections between widely dispersed ethnic groups and races.

Responder

Non-responder

Toxic responder (adverse drug rxn)

SNP

1/2M

siRNA

短片段雙鏈核糖核酸分子 (small interfering RNA molecule)

一些短片段的雙鏈 RNA 能夠使特定的 mRNA降解 (messenger RNA degradation)，藉此阻斷此基因的表現 (block gene expression)，此一作用機制稱為 RNA 干擾現象 (RNA Interference，簡稱為 RNAi)。

siRNA 分子能夠阻斷基因表現的機制，是因為它以與序列同源互補的 mRNA為標靶，降解特定的 mRNA。

RNAi

RNA干擾現象 (RNA interference ,簡稱RNAi)

dsRNA會被一種名為 Dicer的酵素分解成許多小片斷。這些小片斷的 RNA 結合到特定序列之 mRNA上，引發 mRNA序列的降解，使 mRNA 無法轉譯蛋白質，造成該特定基因 “沈默 (gene silencing)”，失去功能。

應用於疾病治療上 可以有效的將病毒與細胞的基因 “沈默” ，抑制病毒的感染與複製能力

經由降解基因的 RNA的方式，可以阻斷基因複製與表現特定蛋白產物。

Applications for siRNA

Basic research– Determining protein function– Easier than a knockout and may be used

for partial knockdowns Clinical research

– Cancer, hypercholesterolemia, infections, developmental defects

microRNA

MicroRNA是一群非常短的 RNA，大約只有二十幾個鹼基，但卻具有影響基因表現的能力。

在抑制基因表現的時候，大多是先變成單股RNA並且與其他蛋白質形成一個複合體， RNA-induced slience complex (RISC)。之後再將與之互補的 mRNA分解。

miRNA最明顯的特徵就是具有一個髮夾的構造(hairpin，即一條 RNA中兩端有互補的序列，中間有一小段不互補的序列，兩端會彼此配對而形成像髮夾一般，有一直段的序列加上一端連接兩邊互補序列的彎曲 ) 。

miRNA Details

Originate from capped & polyadenylated full length precursors (pri-miRNA)

Hairpin precursor ~70 nt (pre-miRNA) Mature miRNA ~22 nt (miRNA) First discovered in 1993 by Victor

Ambros at Harvard (lin-4)

Another View

Microprocessor Complex

What is the Difference between miRNA and siRNA?

Function of both species is regulation of gene expression

Difference is in where they originate siRNA originates with dsRNA siRNA is most commonly a response to foreign RNA

(usually viral) and is often 100% complementary to the target

miRNA originates with ssRNA that forms a hairpin secondary structure

miRNA regulates post-transcriptional gene expression and is often not 100% complementary to the target

Chapter 10 Lipids

1. 每一公克脂肪可以產生九大卡熱量﹐是濃度最高 的能量來源﹐2. 脂肪也是人體儲存能量最主要的方式 3. 脂肪像一層軟墊一樣﹐支持﹑保護神經及內臟器官﹐使它們不容易受到外來撞擊的傷害 .

4. 並且有助於維持體溫。

5. 脂肪也有促進脂溶性維生素（維生素 A D E﹑ ﹑K﹑ ）吸收的效果﹐如果脂肪攝取不足﹐也可能造

成脂溶性維生素缺乏。

6. 脂肪對於人類正常的生長﹑細胞膜的完整﹑荷爾蒙的合成都很重要

脂肪

反型脂肪

天然脂肪酸都是順型 (Cis-)的，而人工合成的脂肪酸是以植物油，經微生物及工業製程加以氫化而成的，稱之為反型脂肪酸 (Trans-fatty acid)。在國內又叫轉化脂肪 (Transfats)、反型脂肪

反型脂肪

反型脂肪的食用可能會影響身體對必須脂肪酸的代謝，致使細胞膜的合成、荷爾蒙的製造產生障礙。

反型脂肪可能也是造成血管硬化的危險因素，它會引起細胞激素的發炎反應產生動脈硬化，破壞血管內皮的一致性，易使動脈硬化腫塊斷裂，發生中風或心肌梗塞的危險。

血脂肪、脂蛋白？ 主要包括膽固醇、三酸甘油酯及磷脂。人體攝入的脂

肪，要能夠在血液中循環，一定要能溶於血中才行，因此利用溶於水的蛋白質來攜帶不溶於水的脂肪。此種血脂肪在血液中與蛋白質結合在一起的結合體，即稱之為「脂蛋白」。

　血液中的脂蛋白主要包括：乳糜小滴、極低密度脂蛋白、中密度脂蛋白、低密度脂蛋白及高密度脂蛋白，此五種脂蛋白的來源、所攜帶的脂肪量與疾病的關係均各有所不同，其中低密度脂蛋白 (LDL)和高密度脂蛋白 (HDL)與血管動脈關係密切，因此做健康檢查時，除了做膽固醇及三酸甘油酯之外，還應該做脂蛋白測定，才能對可能發生的疾病有更正確的預測。

Lipids:

1. Compounds composed largely of reduced carbons (low oxidation state) and exhibiting low water solubility.

2. The longer the fatty acyl chain the fewer the double bonds, the lower is the solubility in water.

3. Lipids may be completely hydrophobic or, if they contain polar groups, amphipathic.

Simple Lipid:

Triacylglycerides (Triacylglycerols): provide stored energy and insulation.

Glycerol esterified with three fatty acids.

Such as: Fats, oils and waxes.

Two advantages to use triacylglycerol as stored fuels rather than glycogen and

starch

1. More reduced. Oxidation can have 2X more energy. Sugar 4kcal/g; lipid 9 kcal/g.

2. Hydrophobic, and unhydrated. It does not have to carry the extra weight of water of hydration that is associated with stored sugar.

Waxes: Esters of long-chain alcohol and long-chain fatty acid;

serve as energy stores and water repellents

Biological Waxes:

Pharmaceutical cosmetic, Lanolin (lamb’s wool), bee wax.

脂肪的種類

脂肪是由甘油及脂肪酸 (Fatty acids) 所組成 typically even of carbons (14 to

24) 飽和脂肪酸

脂肪酸的分子結構為單鍵碳，如 -C-C-C-

不飽和脂肪酸

單元性不飽和脂肪酸 ,一個雙鍵碳，如 -C-C＝ C-C-

多元性不飽和脂肪酸

兩個或兩個以上的雙鍵碳，如 -C＝ C-C＝ C-

Nomenclature

Systematic: e.g., Octadecanoic acid: 18-carbons long

Common names: Stearic acid (硬脂酸 )

Short hand notation:

18:0 -18 carbons long with no double bonds

18:1(Δ9) -18 carbons long with one double bond starting at carbon 9

月桂冠

肉荳蔻

花生酸

亞麻油酸

花生四烯酸

人類血液中的脂質有四種，其中主要的兩種是

1.膽固醇和

2.三酸甘油脂（中性脂肪），

另外還有兩種就是

1.磷脂質和

2.游離脂肪酸。

1 亞麻油酸（ linoleic acid）及次亞麻油酸（ linolenic acid）是人體無法自行製造 ,必需 從飲食取得﹐稱為必需脂肪酸。2.缺乏必需脂肪酸會造成生長遲緩﹐皮膚發炎等疾病。

3. 黃豆油﹑玉米油﹑紅花籽油﹑葵花子油﹑麻油都是必需脂肪酸的良好來源

4. 有些廣告打出的「維他命 F」﹐就是指這兩種脂肪酸

5.To synthesize arachidonic acid -precursor to eicosanoids (Prostaglandins)

必需脂肪酸

血中膽固醇和三酸甘油脂長時間升高時，會引起血管的硬化，而血管硬化後更會引起高血壓、心臟病（心肌梗塞）、腦血管障礙（出血、梗塞）及腎臟衰竭等疾病

脂蛋白因其比重的不同而可分為五種

1.乳糜微滴 (chylomicron)

2.極低密度脂蛋白 (very low density lipoprotein, VLDL)

3.低密度脂蛋白 (low density lipoprotein, LDL)

4.中間密度脂蛋白 (intermediate density lipoprotein, IDL)

5.高密度脂蛋白 (high density lipoprotein, HDL)

2/2M

Complex Lipids:membrane lipidPhospholipids:

Glycerophospholipids (Glycerol+2FA

+PO4)

Sphingolipids (Sphingosine+FA+PO4)

Glycolipids:

Sphingolipids (Sphingosine+FA+sugar)

Galactolipids (Glycerol+2FA+sugar)

A class of phospholipids; also called phosphoglycerides. Polar group esterfied to phosphate of phosphatidic acid (Fig 10-8)

*1,2-Diacylglycerol with phosphate ester at carbon 3

*Phosphatidic acid: Glycerol-3-phosphate with fatty acids esterified to C1 and C2

C1: fatty acid typically saturated

C2: fatty acid typically unsaturated

Glycerophospholipids:

Some phospholipids have ether-linked fatty acids

Ether glycerophospholipids:

Ether linkage at carbon 1

Alkyl group esterfied to carbon 2

Plasmalogen: Ether glycerophospholipids with unsaturated alkyl group. (Fig 10-9)

Platelet-activating factor :

Plasmalogen:

1. Half of heart phospholipid are Plasmalogen.

2. The functional significance is unknown.

Platelet-activating factor :

• Cause platelet aggregation

• Potent mediator in inflammation, allergic response and shock

• Molecular signal

• Release from leukocytes

(D) 5. 支氣管哮喘症 (Bronchial asthma) 的早期變化約在暴露於過敏原後 30 分鐘內發生，其致病機轉與下列物質有關，何者除外 ?

A.Histamine B.Leukotrienes C.Prostaglandins D.Dopamine E.PAF(platelet-activating factor)

Sphingolipids:

Backbone of sphingosine: 18-Carbon amino alcohol with C-C trans double bond. Sphingolipids are derivatives of Sphingosine. (Fig 10-12)

Sphingomyelins:

1. In plasma membrane

2. A membranous sheath in some neurons

Sphingolipids:

當 Sphingolipids 缺乏時，成熟精子的數目會顯著的下降，老鼠受孕的可能就大打折扣了 . Proceedings of National Academy of Sciences 2002;10:1073

Glycosphingolipids at cell surfaces are sites of biological recognition: (Fig 10-14)

Blood A, B, O.

 

Phospholipids and Sphingolipids are degraded in lysosomes

-Genetic Defect Diseases: Box10-2; abnormal accumulations of membrane lipids.

Phospholipid &Sphingolipids are degraded in lysosomes

Niemann-PicK disease

1. Sphingomyelin accumulates in brain, spleen and liver

2. Mental retardation, early death

Tay-Sachs disease

1. Ganglioside GM2 accumulates in brain, spleen.

2. Progressive retardation in development, paralysis, blindness, death 3, or 4y.

尼曼皮克氏症(Niemann-Pick disease)

尼曼皮克氏症是一種罕見的神經退化性疾病，或稱為鞘髓磷脂儲積症，起因於體內無法將膽固醇運送到正確的位置，產生醣酯堆積而導致神經髓鞘受損

發病年齡通常在學齡期前，但也有嬰兒期就發病或成年才發病的晚發型 症狀以肝脾腫大為主，但是神經系統病變是最主要問題，包括眼球無法垂直轉動，肢體張力過強、協調力平衡力差，導致行動笨拙而變得不靈光，說話緩慢，發音也不清楚。他們的反應力、記憶力、協調能力會逐漸退化，飲食時也會發生吞嚥困難、甚至喝水嗆到的問題，走路平衡感變差容易跌倒。到後期肌肉無法控制會導致長時間臥床，甚至死亡。發育遲緩、學習障礙與智能不足使患者的一般表現差於正常人

對這樣一種嚴重、致死性的遺傳疾病而言，各醫學中心每年都會見到幾個病例。但是在治療方面，一直到最近為止，並沒有有效的治療。過去有人嘗試骨髓移植，卻無法阻滯疾病持續惡化。

Tay-Sachs disease

罹患此症的嬰兒因缺乏一種己醣胺酵素 A(Hexosaminidase A)，而此酵素的重要功能在於與神經節甘脂 (Gangliosides)代謝有關，特別是 GM2,；這些 GM2物質會聚集並逐漸毀壞腦部與神經細胞，直到中樞神經系統功能完全停止。神經節甘脂 (Gangliosides)是存在於神經細胞膜上的一種重要的醣脂，正常情況下會被溶小體中的 β -galactosidase酵素所分解。如果 β-galactosidase缺乏或酵素功能不良，導致 GM1神經節糖堆積在神經細胞或其他內臟組織中，便會產生臨床症狀。為體染色體隱性遺傳 。Tay-Sachs disease症患者的 hexosaminidase A酵素活性缺乏。而 Sandoff disease症患者為 hexosaminidase A 及B 兩種酵素活性都缺乏。典型的嬰兒期患者，此症為一致命性疾病； 臨床表徵主要以多變化的神經學症狀為主，症狀包括步態不穩、四肢僵硬、近端肌肉萎縮與構音困難。

高雪氏症 (Gaucher's disease)

Gaucher’s Disease- rare inherited disease; deficiency or lack of an enzyme called Glucocereborsidase

Results in an accumulation of glucocerebroside within cells in various body tissues ( spleen, liver, bone marrow, and skeleton)

Severity of the disease can vary

Symptoms

Bone pain

Victim bruises easily and there are many fractures in their bones

Difficulty walking Blood clotting

Symptoms (continued)

Muscle weakness

Poor coordination

Seizures

Prevention

No real prevention

Genetic Counseling is recommended for parents with a family history of the disease

Treatments

No CURE Enzyme replacement therapy-injections of

the enzyme Result: decrease liver and spleen size

reduce skeletal abnormalities

restores normal growth & development

restores well being of the patient

Tay-Sachs disease, Ganglioside deposites in lysosomes

Fabry Disease

Progressive Multiple organ systems Morbidity

• Cardiac complications

• Stroke

• Renal failure

• Decreased lifespan

Fabry Disease X-linked inborn error of metabolism

Deficient -Galactosidase A (-GAL A) enzyme activity

Progressive globotriaosylceramide (GL-3) accumulation

– multiple cell types and tissues -- end organ impairment

Metabolic Pathway

Sterols have four fused carbon rings

Cholesterol

Steroid hormones

Androgens:Testosterone

Estrogens: Estradiol

Progestins:Progesterone

Glucocorticoids: Cortisol

Mineralocorticoids

Bile salts: Cholic acid and deoxycholic acid

 

Lipids as signals, cofactors, and pigments

Membrane lipid: 5-10% of cells

Storage lipids: >80% of adipocyte.

Phosphatidylinositols act as intracellular signals

Fig 11-15. 1,4,5-triphosphate (IP3)—Ca++ release.

Ceramids and sphingomyelin are potent regulators of protein kinases.

Eicosanoids carry messages to nearby cells

paracrine hormone involved in:

reproductive function

inflammation function

formation blood clot, regulation blood pressure derived from 20-carbon polyunsaturated fatty acid arachidonic acids 20:4 (Δ5,8,11,14).

Arachidonic acids 20:4 (Δ5,8,11,14)

Eicosanoids, 3 classes

A. Prostaglandins:

1 regulate cAMP.

2 stimulate contraction of smooth muscle of uterus, pain for MC and Labor.

3 affect blood flow.

4 affect wake-sleep cycle.

5 elevate body temp.

B. Thromboxanes:

1. produced by platelets. Act in formation of blood clots, reduction blood flow.

2. NSAIDs (non-steroidal anti- inflammatory drugs): aspirin, ibuprofen, acetaminophen.

C. Leukotrienes:

Derived from leukocytes. Induces contraction of muscle lining of the lung—asthmatic attacks or anaphylactic shock.

Prednisone-anti-asthmatic, anti-rheumatoid arthritis drugs.

Do cosa hexaenoic acid ( 二十二碳六烯酸 , DHA)含有 6 個雙鍵的多元不飽和脂肪酸，屬於必需脂肪酸，DHA為大腦與神經組織中細胞膜的主要成分，在人體內自行合成的能力相當低，所以必須從外界攝取來補充。

DHA 是一種增加細胞膜流動性與滲透性的重要必需脂肪酸。此一現象能讓細胞感應器發揮適當功能，調節營養素和廢物在細胞與胞器內外的進出 .

DHA 是細胞膜中最長也最高度多元不飽和的脂肪酸，以神經細胞膜、視網膜和睪丸中的濃度最高。

Steroid hormones carry messages between tissues

Vitamins A and D are Hormone precursors (Fig 11-8)-Rickets

** Vitamins E and K and the lipid Quinones are oxidation-reduction cofactors

Vit E:

*Tocopherols-antioxidants.

*Deficiency: scaly skin, muscular weakness, vasting, sterility.

Vit K:

*Blood clotting.

* Warfarin: inhibit the formation of prothrombin. Anticoagulation drug; poisonous to rats.

 

Chapter 11

Biological membranes and transport

Membrane functions:

Boundary for cell and organelles

Surface on which reactions can occur

Regulation of material flux through membrane proteins

Signal transduction interface

Membrane components:

Lipids: Amphipathic molecules arranged as bilayers with polar groups out and nonpolar groups in

Membrane Proteins:

Surface associated with or embedded in bilayer

Functions: Transport, receptors, enzymes

Two types: Peripheral and integral

Plasma membrane:

*Delimits cell

*Excludes and retains certain ions and molecules

*Cell locomotion

*Reproduction

*Signal transduction

*Interactions with other cells or extracellular matrix

*Lipid bilayer: Two lipid monolayers with hydrophobic surfaces face to face

Cell surface polysaccharides:

involved in modulating response of cells to other cells or to extracellular matrix,

Interactions involve glycoproteins and proteoglycans

Fluid Mosaic Model

Singer and Nicholson, 1972

Phospholipid bilayer forming fluid matrix

Two classes of membrane proteins

1, Peripheral (extrinsic) proteins

Associated with bilayer surface via ionic interactions and H bonds. Extractable with high salt or agents that disrupt H bonds (urea).

2, Integral (intrinsic) proteins

Associate with hydrophobic bilayer interior via hydrophobic interactions. Extractable with detergents .

Depending on the conditions and the nature of the lipid, when amphipathic lipids are mixed with water

1. Micelles

2. Bilayer

3. Vesicle or liposome

Membrane lipids are in constant motion

1. Lateral diffusion

2. Transbilayer diffusion

Membrane mobility

Protein

Frye and Edidin, 1970: Lateral movement of membrane proteins following fusion of mouse and human cells

Lateral movement may be impeded by interactions with cytoskeleton

Lipids

Rapid lateral movement, slow transverse movement

Lipid-linked membrane proteins:

Cys, Ser, Gly

Sugar-linked amino acid

Ser, Thr, Asn

Four types of intergral p in cell-cell interaction

Cell-cell interaction

1. Integrins: heterodimeric proteins Binding site: Arg-Gly-Asp (RGD)2. Cadherin: homophilic3. N-CAM: hetero or homo interaction. 4. Selectins: Ca++ domain, bind poly- saccharides.

Caveolin

1. Associate with cholesterol, contain 3 fatty acyl moieties

2. Forming caveola

3. Membrane trafficking

4. Transduction of signal

Membrane fusion is central to many biological processes

1/3M

Membmne transport: Three types

#Passive diffusion:

Entropically driven process: Molecules move down a concentration gradient

Rate depends on concentration gradient and lipid solubility

#Facilitated diffusion or passive transport:

Entropically driven process as in passive diffusion

Involves integral membrane protein

Rate depends on concentration but is saturatable & specificity

Specificity and affinity due to protein/transported molecule interaction

Transporter (integral protein)

Symport: Ion and substance move in same direction

Antiport: Ion and substance move in opposite directions

Uniport: Only one substance move across

*Transporters or permeases

Examples: 1.Glucose transporter 2.Anion transport (exchanger for HCO3-) system and 3. Aquaporins

1. Glucose transporter:RBC

RBC band 4.5: 45 kD Protein -functional as trimer

GluT1: specific for D-glucose.

GluT2: transport glucose out of hepatocyte

5mM Glucose at body. 50000X greater than diffusion rate.

Glu T1

GluT1:

glucose transporter

High rate

Saturability

specificity

2. Anion transport (exchanger) system:

RBC band 3: 95 kD Protein: chloride and Bicarbonate system. Fig 12-28.

Chloride-bicarbonate cotransportHCO3- is much more soluble in blood plasma than is CO2

3. Aquaporins (AQPs): integral proteins, provide channels for rapid movement of H2O.

AQP-1

Defective glucose and H2O transport in 2 forms of Diabetes

Type I (Juvenile onset)

Type II (Adult onset)

Diabetes insipidus: a genetic defect in AQP-2 leads to impaired water reasorption.

ADH: antidiuretic hormone, regulates AQP-2

2/3M

Active transport: Energy driven process: Fig 11-35.

Primary active transport: Energy Sources

A, ATP hydrolysis (most common)

B, Light energy

Secondary active transport:

Energy is ion gradient formed by some other process

Active Transport-Driven by ATP

4 types of transport ATPases:

A. P-type ATPases:

*Na+,K+ ATPase (Sodium PUMP, Fig 11-36)

120 kD alpha-subunit, 35 kD beta-subunit

3 Na+ out, 2 K+ in per ATP hydrolyze : Electrogenic

Ouabain, digitalis: Cardiac glycoside that inhibits sodium pump

毛地黃類毒素 (CARDIAC GLYCOSIDE)

夾竹桃 (Oleader) 玲蘭 (Lily of the valley)

毛地黃 (Foxglove)

1.毛地黃素是用來治療嚴重心臟病的藥物，是由毛地黃提煉出來的物質。但是在野外如果不小心誤食了這類的植物，反而會造成心臟的嚴重問題，嚴重的會有昏厥、驚攣，甚至呼吸心跳停止的現象。

2.毛地黃為重要的強心藥，可興奮心肌，增強心肌的收縮力，改善血液循環。

P-type ATPase,

phosphorylation as part of transport cycle.

Sensitive to Vanadate

Sodium pump (Na+K+ ATPase)

Neuron communication

Regulation of cell shape and volume

Transport amino acids, sugar, nucleotides..

Consumption 25% of metabolic energy at rest (70% at neuron)

Jens Skou 1957

1/3C

Monarch Viceroy

Ouabain, digitalis (foxglove plant):

A potent and specific sodium pump (Na+K+ ATPase) inhibitor.

Inhibition of Na+ efflux by digitalishigher Na+ conc. In cellsactivating Na+-Ca+ antiporter in cardiac muscle Ca+ increase concmuscle contraction.

Calcium ATPase

2 Ca+2 out of cytoplasm per ATP hydrolyzed

Restores/maintains low cytoplasmic calcium

Store Ca+2 at ER lumen

H+, K+-ATPase

1 H+out, K+ in per ATP hydrolyzed

1K+, Cl- cotransporter out

Net : HCl produce.

Gastric enzyme: ΔpH largest gradient known (PH 0.8~16.67.4)

B. V-types ATPases:

Vacuolar ATPases: Pump H+ in number of vacuoles and cells (lysosomal, endosomal secretory vesicle).

(C) F-type

ATPase:

ATP synthase

High H+

C. F-types ATPases: ATP synthase

energy-conserving reactions (ATP) in mitochondria, bacteria, chloroplast.

D. ABC transporter:

Multidrug resistance transporter: in malignant cells transported by ATPases

A defective ion (Cl-) channel cause Cystic Fibrosis

High NaCl

Cystic Fibrosis 纖維囊腫

致病基因位於第七染色體的長臂 (q arm):q21～ q31之間。全長約 25萬個鹼基長 (250kb).

一種遺傳疾病，其特徵在出生後不久即會表現。病人肺內黏液很多，肺表皮細胞死去，長成囊腫，呼吸困難。原因是基因異常，無法控制氯離子通過細胞膜。纖維性囊腫造成極高的幼兒死亡率，但隨著醫療的進步，病患通常可以活到 20歲以後。

Driven by Light

Bacteriorhodopsin: Proton pump

Halorhodopsin: Chloride pump

Secondary active transport:

Na+ or K+ Coupled movement of amino acids or sugars (Fig 11-44)

Pump glucose inward driven by high extracellular [Na+] until about 30,000 times that in the intestine

Valinomycin (K+) and monensin (Na+)—antibiotics,

ionophores, compound that shuttle ion across membrane.

Kill cells by disrupting secondary transport processes.

Valinomycin

Ion Channels are different from ion transporters

1. Orders of magnitude greater than transporter.~107~108 ions per channel per second.

2. Not saturable

3. Gated opened or closed in response to signals.