2025年生物化學入門指南探索生命科學的基石

BIO107IntroductiontoBiochemistryGuoxiaHan

Lct.l

Questions

Chemicalstructure,interaction,extractenergy,storeandtransmittheinformation,chemical

changes,controlled

3principleareas

StructuralChemistry,

Metabolism,

Storage,transmission,andexpressionofgeneticinformation

Lct2,ChO3

AminoAcids,Peptides,andProteins

AminoAcids

COO-Isomers

Structuralisomers構(gòu)造異構(gòu)

體

H3N—C—HCis-transisomers順反異構(gòu)

體

REn-antionmers光學異構(gòu)

山HM口4-/.3

Stereoisomers空間異構(gòu)體（Glycer-aldehyde甘油醛、Ala丙氨酸）

,CHOCHO

i

-H一OH

CHQH

L-Glyceraldehydeo-Glyceraldehyde

COO-COO-

?1

H3N-￡-HH-C-

CH3CH,

L-Alanineo-Alanine

蛋白質(zhì)中出現(xiàn)的氨基酸都是Lisomers

>AminoAcidsClassifiedbyRGroups

Nonpolar,aliphaticRgroups

COO"COO-coo-coo-非極性脂肪基

?1?1l/H?1R

HN—C—HHN—C-HHN—C—H

333Glycine甘氨酸Gly,G

11HNCH1

HCH22CH

3AlanineAla,A

HCCH/\丙氨酸

22CHCH

33Proline脯氨酸Pro.P

GlycineAlanineProlineValine

Valine繳氨酸Val,V

COO'COO-coo-

???Leucine亮氨酸Leu,L

n3nvriH3N—C—HH3N—C—H

w3Isoleucine異亮氨酸

3He,I

/火CH2

AA…L'TccRATH

CH3CH3CH3s

1

CH3

LeucineIsoleucineMethionine

Polar,unchargedRgroups

coo-coo-coo-

?I.1

H3N—C-HH3N—C-HH3N—C—H

CH20HH—C—OH

I臚

CHSH

3極性不帶電R基

SerineThreonineCysteine

Serine絲氨酸Sens

coo-coo-

?IThreonine蘇氨酸Thr,T

HN—C—HHN—c—H

33Cysleine半胱氨酸

段Cys,C

心Asparagine天冬酰胺

/%Asn.N

H2NO

AsparagineGlutamine

PositivelychargedRgroups

coo-coo

正電R基

Lysine賴氨酸Lys,K

Arginine精氨酸Arg,R

Histidine組氨酸His,H

Guanidino服基

Imidazole咪嚶

C=NH,

NH2

LysineArginineHistidine

AromaticRgroups

芳香族R基

Phenylalainine苯丙氨酸Phe,F,

非極性

Tyrosine絡氨酸Tyr,Y,極性無電

Tryptophan色氨酸Trp,W,非極性

Rotate

ring

r1.I-:nainn-rr

PhenylalanineTyrosineTryptophan

1、非極性氨基酸（疏水氨基酸）8種

丙氨酸（Ala）綴氨酸（Vai）亮氨酸（Leu）異亮氨酸（He）脯氨酸（Pro）苯丙氨酸

（Phe）

色氨酸（Trp）蛋氨酸（Met）

2、極性氨基酸（親水氨基酸）：

1）極性不帶電荷：7種

甘氨酸（Gly）絲氨酸（Ser）蘇氨酸（Thr）半胱氨酸（Cys）

酪氨酸（Tyr）天冬酰胺（Asn）谷氨酰胺（Gin）

2）極性帶正電荷的氨基酸（堿性氨基酸）3種賴氨酸（Lys）精氨酸（Arg）組氨酸

（His）

3）極性帶負電荷的氨基酸（酸性氨基酸）2種天冬氨酸（Asp）谷氨酸（Glu）

AlanineAlaAnonpolarNeutral1.8

GlycineGlyGnonpolarNeutral-0.4

IsoleucineHeInonpolarNeutral4.5

LeucineLeuLnonpolarNeutral3.8

MethionineMetMnonpolarNeutral1.9

PhenylalaninePheFnonpolarNeutral2.8257.206,1880.2,9.3,60.0

ProlineProPnonpolarNeutral-1.6

TryptophanTrpWnonpolarNeutral-0.9280,2195.6,47.0

ValineVaiVnonpolarNeutral4.2

positive(10%)

HistidineHisHPolar-3.22115.9

neutral(90%)

ArginineArgRPolarPositive-4.5

LysineLysKPolarPositive-3.9

AsparagineAsnNPolarNeutral-3.5

CysteineCysCPolarNeutral2.52500.3

GlutamineGinQPolarNeutral-3.5

SerineSerSPolarNeutral-0.8

ThreonineTb.rTPolarNeutral-0.7

TyrosineTyrYPolarNeutral-1.3274.222,1931.4,8.0,48.0

AsparticacidAspDPolarNegative-3.5

Glutamicacid（；h：EPolarNegative-3.5

Reversibleformationofdisulfidebond

兩個半胱氨酸的端基形成二硫鍵

H3N

Cysteine

Cysteine

Absorptionofultravioletlightbyaromaticaminoacids

色氨酸、絡氨酸的紫外線吸取圖

f,0lW

?rXZW'?1*.h”

Lambert-BeerLaw

log(iyi)=8CL

Uncommonaminoacidsalsohaveimportantfunctions

aPo*d'fp

?NH

ptonlcalval.3

y<arboxygluUmirte

■■*anHE/OO

4*Hydroxyprolin€CH

片“刈

-CH>-CH3〉CHTCH、KM》,

OH

SHydroxytysine

2網(wǎng)2

整~2

6-N-MethyllysineDesmosine

ResiduescoatedbyModificatioitofCOMMonresiduesalreadyincotpora^dintoapolypeptide

HSe-CH,—CH—COO*

rare,dufingpeQ力?300additionalaminoadds

*NHsyniNsisratherthanceaied

thavebeenfoundincells

S?leno<yst?inethmughapostsy泄論如modification

植物細胞壁膠原質(zhì)collagen,膠原質(zhì)co壁gen,肌漿球蛋白myosin,prothrobim,elastin彈性蛋白

細胞中壞發(fā)現(xiàn)了其他?300種氨基酸

這些殘基residue通過調(diào)整已經(jīng)組合成多肽的氨基酸的常見殘基而形成

Reversibleaminoacidmodificationsinvolvedinregulationofproteinactivity

氨基酸的可逆修飾控制蛋白質(zhì)的活動

-o-r—0—CMi-^w-coo

產(chǎn)一!—?

-0p-o-CHTH-COQ-

)E.GluUmat#riMfhyl<st?r

H>o9pH<Mhrvonene

1-fM-COO-

PhMfriMtyrOMn?

-NH<CH)>CHa-O?1-CH-COO*

-CH,-O?i-CM,-CH；-,"TOO-

Aminoacidscanactasacidsandbases

氨基酸的酸堿兩性

HH

I

R-C-COO*—COO-+H+R-C-C二0F?。

HNOHH,N+O-

*NH.NH22

ZwitterionNonk)nkformZwitterionicform

Macid

HHR-C—COO-+H*—,*

?NH,

*NHj?NH,Zwittarion

Zwitterion?sbase

asbase

Non-ionicform非離子態(tài)；Zwitter-ionicform兩性離子態(tài);Amphoteric兩性的

對Gly的titration滴定

Isoelectricpoint(orisoelectricpH)

pl=%(p/+pfc2)=%(2.34?9,60)=5.97

對比Gly滴定與乙酸、甲按滴定成果

P*a24681012

Methyl-substituted

carboxyland

aminogroupsCH,-COOH

H*

AceticacidMethylamine

Th?normalpKaforaThenormalpK.foran

carboxylgroupisabout4.8.aminogroupisabout10.6.

Carboxyland

aminogroupsH-i-COOH&

inglycine

HH*

<t*Aminoacid(glycine)u-Aminoocid(glycine)

叱?234叫片9g

RepulsionbetweentheaminoElect/onefatlveoxygenatoms

groupandthtdefxartingprotoninthecarboxylgrouppullelectrons

lowerstheforthecarboxylawayfroentheaminogroup,

group,andoppositelychargedloweringHspK.,

groupslowerthepKabystabi"

lizicg1h<zwitterion.

Aminoacidsdifferintheiracid-baseproperties

of-COOHgroup:1.8?2.4

R基donotionize的氨基酸

of一NHjgroup:8.8?11.0

IonizableR基的氨基酸e

三個ionization電離階段f3個p口值

Peptidesarechainsofaminoacids

RHR2

H3N—CH—C—OH+H-N-CH-COO'F

IICNjOMHHHCM2HCKtH

OHJM-C--^-H-^-COOT

型檢附sHq］卜電。condensationHOH0M0HON

AminoCarboxyl-

terminal<ndterminalend

R1HR2Serylglycityros\1alanylleudne

?IIIor

H3N—CH-C—N-CH-COO'Ser-GIy-Tyr-ZMa-Leu

Hor

°SGYAL

Hydrolysis&CondensationPeptidebond肽鍵Di-peptide二肽Penta-peptide三肽

寡肽、低聚肽oligo-peptidepolypeptide多肽

Multi-subunitproteins有兩個或更多的多肽鏈(noncovalently)結(jié)合

假如至少有兩條鏈是iden:ical完全,致的-則這個蛋白質(zhì)是oligomeric低聚物的，

而這些同樣的units(包括一種多種鏈)被稱為protomers原聚體

自然產(chǎn)生的蛋白質(zhì)的residues殘基一般少「2,000

Polypeptides有獨特的氨基酸構(gòu)成

某些蛋白質(zhì)包括不是氨基酸的chemicalgroups（conjugatedproteins并合的蛋白）

TABLE3-4ConjugatedProteins

CfassProsthebcgroucExample

LipoproteinsLipids/3t-Lipoprotein

ofblood

GlycoproteinsCarbohydratesImmunoglobulinG

PhosphoprotensPfiosphate?oupsCaseinofmilk

HemoproteinsHeme(ironporfhynn)Hemoglobin

FlBvoprotemsRavinnucleotidesSuccinate

dehydrogenase

MetalloproteinsIronFemtin

ZincAlcohol

dehydrogenase

CalciumCalmodulin

MolybdenumDlnttrogenase

CopperPlastocyanin

這些非氨基酸的部分一般被稱為prostheticgroup輔基

>ProteinSeparationandPurification

ColumnChromatography色層分離法

Crudeextract粗提取今今今Fractionation分播

Ion-ExchangeChromatography

離子互換色層分離

?Largenetpositivecharge

。Netpositivecharge

cNetnegativecharge

?Largenetnegativecharge

包括負電官能團的聚合物球珠

蛋白質(zhì)混合物與陽離子互換劑加入試管

蛋白質(zhì)下滲的速度由他們在目前ph下的凈電量決定。

陽離子互換劑使得負凈電量高的蛋白質(zhì)移動更快被更

早洗提elute出來。

Proteinmixtureisadded

tocolumncontaining

cationexchangers.

Proteinsmovethroughthecolumnatratesdeterminedbytheir

netchargeatthepHbeingused.Withcationexchangers,proteins

withamorenegativenetchargemovefasterandeluteearlier.

ExclusionChromatography(gelfiltration)

排斥色層分離(膠體分離)

多孔滲水的聚合物球

蛋白質(zhì)混合物與cross-linked聚合物加入試管

蛋白質(zhì)分子根據(jù)大〃環(huán)一樣而被分離

更大的分子更輕易通過，出目前更早的（分）儲（部）分

fraction

Proteinmoleculesseparate

bysize;largermolecules

passmorefreely,appearingn

intheearlierfractions.123456

AffinityChromatography

同類色層分離

Proteinof

interest

ligand

蛋白質(zhì)混合物

惴/爆

MixtureSolution蛋白質(zhì)混合物與結(jié)合苕配體ligand的聚合物加入試

七

ofproteinsofligand管，

；配體可以與想要的蛋白質(zhì)結(jié)合

Proteinmixtureb

addedtocolumn雜質(zhì)雷白被沖出試管

containinga了

polymer-bound配體熱溶液

ligandspecificfor

proteinofinterest.

4)

UnwantedproteinsProteinofinterest

arewashedthroughiselutedbyligand

column.solution.

TABLE3-5APurificationTableforaHypotheticalEnzymeI

FractionvolumeToialoroteinActMtySpecificactivity

Procedureorstep(ml)(mg)(units)(unlts/ngj

1.Crudecellularextract40010000100,00010

2.Precipitationwithammoniumsulfate2803.00096,00032

翼

3.Ion-exchangechromatography即40080,0002C0

4.Size-exclusionchromatography10060,0006CO

5.Affinitychromatography345,00015.0CO

Note:AlldatareoresentthestatusofttesampleaftermedesignatedprocedurehasbeenearnedoutActivityandspecificactivityarede

finedonpage94..

Activityfumts，表達一份溶液中酶的總單彳立數(shù)量

Specificactivity(units/mg)表達每毫克總蛋白質(zhì)酹的單位置；酶的純度

1.0unitofenzymeactivity=在25℃和最佳測量條件下,每分鐘引起1.0nmol底物轉(zhuǎn)化的酶

的數(shù)展(amountofenzymecausingthetransformationof1.0pmolofsubstrateperminuteat

25℃underoptimalconditionsofmeasurement)

Electrophoresis電泳

SDS-polyaeryIamidegel

Crosslinkedpolymer

polyacrylamide

letsasamolecularsie^e,sbwingthemigration

proteinsappradmateJ^inproportiontotheir

harge-to-massratio.

Relativemigration

IsoelectricfocusingTwo-dimensionalelectrophoresis

TABLE3-6TheteoeloctrkPointe

ofSomeProteins

A-oewpl

<1.0

照altxiE力4.6

Seamalbunvn4.9

Urease5.0

/Racto^obuln5.2

Hemogjotxn68

MyogioDtn7.0

Chymotrypsinogen9.5

C?tocfiron>ec10.7

Lysaome11.0

Thereareseverallevelsofproteinstructure

PrimarySecondaryTertiaryQuaternary

structurestructurestructurestructure

AminoacidaHelixPolypeptidechainAssembledsubunits

residuesf.

1氨基酸殘基的線性序列

（氨基酸殘基residues,包括二硫鍵）

2局部穩(wěn)定的氨基酸殘基序列所形成反復出現(xiàn)的構(gòu)造模型，包括a-螺旋，B?折疊，B?轉(zhuǎn)

角，無規(guī)卷曲四種構(gòu)造

3在二級構(gòu)造基礎上深入折疊成緊密的三維形式

（多肽鏈，多種3d折疊后的多肽）

4由兩個或多種多肽鏈（亞基）互相作用形成的空間構(gòu)造。

（由蛋白質(zhì)亞基構(gòu)造形成為多于一條多肽鏈的蛋白質(zhì)分子的空間排列）

（組合在一起的subunit,形成Multi-subunitProtein,Arrangementisspaceofpolypeptide

subunits。）

Determinationofaminoaddsequence

Theaminoacidsequencesofmillionsofproteinshavebeendetermined.

數(shù)百萬種蛋白質(zhì)的氨基酸序列已經(jīng)被確定

Shortpolypeptidesaresequencedusingautomatedprocedures

Sanger'smethod

測定amino-terminal殘基

TheEdmandegradationprocedure

降級過程

由一種測序儀進行

Largeproteinsmustbesequencedinsmallerfragment

打斷disulfidebond

Osulfidebond

C-o

Acetylated

cysteine

扯破Cleaving多肽鏈

TABLE3-7RieSpecificityofSomeCommon

MethodsforFragrncntingPolypeptideChains

Reagentfdio<a0cdisourcerOeavo^powits1

Trypsinlys.Arg(C)

(bovinepancreas)

SubmaxiHantsprotease〃(C)

(mousesuomanllary0and)

Clvnotr>psinPhe.gM(C)

(bovinepancreas)

StapftyJooocct/saureusV8proteaseAsp.Glu(C)

(bacte?iumS.aureus>

AspWproteaseAsp.Glu(N)

(bactenumPseudomonas^agi)

RopjmPh?,Trp.Tyr(N)

dxxeinestomach)

LndopcoteioaseLysClys(Cl

(bacteriumLysobacter

enzymogeres)

Cy<aix)eentxonudeM?(C)

?AJm即THflaacxcym9tobmrrw^am⑷AmMMiUbM

cawnercMsources.

咯5加埠gprtflMiy但W6OOHlortfwBON統(tǒng)or

0m：pwodetmSoccirsonrthetthecabonyl(C)orttie

aniro?N)MedZgc“edsmnoaodmMLn.

某些proteases蛋白酶只把破臨近特殊氨基酸殘基的肽鍵

Orderingthepeptidefragments

ProcedureResultConclusion

h心。lyt?:ASH2I*11Polypeptidehas38

IVHWIQMH9C2J_1$2dminoaddresidues.Tryp?

D4IK~T1JrwillckavethreUes

t2J4V1(atoo*R(ArgIandtwo

FiIMdv2

PolypeptideK(Ly$Htogivefour

G3PI

trnnts.Cy?no9?nbromide

willclMveattwo

M(M#t)togiveth，”

reectwHtiFDNfe;hydrolyM；fragments,

2(4-0inHroptwnylglut?matvE(Glu>杯imino-

rXuc?detectedtecminalresidue.

<Skuffid?

tendsIlfp/eseotl

?

deeve.0；?CASMALIK，力p&cdataminoterminus

5?URItb?9lnswithE(Glu).

?EGAAYHOFEPIDPR

()pU<Xatcarboxylterminus

DCVMSO

o-becauseitdoesnotendwith

YLIACGPM1KR(Aro)orK(Lys).

(Move“MM9M

bwnitff；tepervftfr”nwntxki)EGAAYHDFEPIOPRGASM4i)ov?rljipswith

￥6―0?g6dm.c

,C2\TKEXVHSD(；And^)Mawing

Mel|C)

XJ)ALNCYLIACGPMthemtob?ordertd.

@?0?

匕JsAYHXM”%同鵬UypMvHSD:由蹙

Aminoacidsequencescanalsobededucedbyothermethods

Aminoacid

sequence(protein)Gin-Tyr-Pro-Thr-lle-Trp

I11111111Ii]

DNAsequence(gene)CAGTATCCTACGATTTGG

■?w.

8d。"

Investigatingproteinswithmassspectrometry

光譜測定法

uilMS3Xecw

-V-*"jL^J

100

I

E75

U5O

W2CS

A

A

e