1-Nucleotides and Nucleic Acids
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Transcript of 1-Nucleotides and Nucleic Acids
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Powerpoint #1: Nucleotides and Nucleic Acids
Chemistry and Biochemistry 153B
DNA, RNA, and Protein Synthesis
Winter 2016 - A!ert Co"rey
UCLA
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1.2) Organizing Principle of the Course=Central Dogma of Molecular Biology
rNPs
DN! "N! Proteintranscription translation
Meta#olites$amino aci%s& 'COO(& etc.)
De noosynthesis
salagesynthesis
Brea*%o+n pro%ucts$+aste pro%ucts& useful meta#olites)
cata#olism%NPs
processing
replication
repair
,ast - ,ectures
/irst -2 ,ectures
Why we study this stuff:0 #1 reason$ “the universe enjoys being observed”* and so the "ni%erse
created creat"res i&e yo" and me 'ho en(oy o!ser%in) the "ni%erse* 0 Other reasons the mechanisms #y +hich genetic information is enco%e%
an% %eco%e% are central to our un%erstan%ing of +ho +e are as human
#eings& an% to our un%erstan%ing of human physiology an% human %isease.
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oet& oet& an% Pratt /igure (
1.) Coalent 3tructure ofNucleic !ci%s
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!) ! purine resi%ueB) !n a%enine resi%ue
C) ! 45 en% resi%ueD) ! %eo6ya%enosine resi%ue7) None of the a#oe
iClic*er 8uestionTo which of the following things is the
red arrow NOT pointing?
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1.9) Nucleoti%e 3tructure
SugarRibose or Deoxyribose
BasePurine or Pyrimidine
Phosphates1, 2, or 3
Nucleosi%e
Nucleoti%e
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1.4) he 3ugars
/ischerPro:ection
C
D(ri#ose,inear al%ehy%e
;(D(ri#ose
$this anomer foun% inri#onucleoti%es an% "N!)
;(D(25(%eo6yri#ose$foun% in %eo6ynucleoti%es
an% DN!)
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1.?) he Bases
he #ases in nucleoti%es aresu#stitute% %eriaties of t+o%ifferent heterocycles.
pyrimidine purine
hey are classical Bronste% #ases& #ecause N1 an% N of pyrimi%ine& an% N1& N& an%N@ of purine hae pairs of non(#on%ing electrons that can #e use% to form coalent #on%s toprotons. pAa5s are in the 9(4 range.
8uestions
1) ill the nitrogens #e protonate% at physiological p'> hy or +hy not2) hy isn5t N of purine #asic
:
: :
:
:
:
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iClic*er 8uestionWhich of the following statements describes the
state of the N1 N! and N" nitrogen atoms in purine
and pyrimidine at physiological pH ? !. hey +ill not #e protonate% #ecause their pAa5s are too high.
B. hey +ill not #e protonate% #ecause their pAa
5s are too lo+.
C. hey +ill all #e protonate%.
D. Only N@ +ill #e protonate%.
7. E %on5t *no+.
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iClic*er 8uestionWhy isnt N$ of the purine ring basic?
!. N is sp hy#ri%ize% an% therefore cannot accept a proton
B. he non(#on%ing electrons on N are %elocalize%
C. Protonation of N +oul% lea% to a loss of resonancesta#ilization
D. Both B an% C
7. !ll of the a#oe
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1.F) Purine an% Pyrimi%ine Deriaties in DN! an% "N!
N
N
N
N
X
X
N
N
X
NH2
O
O
O
O
O
CH3
N
NN
N
H
H
X
N
NN
N
X
NH2
H2N
H
O
C
G
!
H
E%iosyncratic nomenclature
12
9
4
?
1
2
9
4? @
F
Bases Nucleosides Nucleotides
Cytosine Cytidine Cytidylic acid
racil ridine ridylic acid
!hymine !hymidine !hymidylic acid
"denine "denosine "denylic acid
#uanine #uanosine #uanylic ac i d
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1.) Base !romaticity
• Planarity of aromatic #ases allo+s efficientstac*ing>. – Because of %elocalize% π or#ital electrons& a%:acently
stac*e% #ases attract one another strongly throughfaora#le in%uce% %ipole interactions.
– his stac*ing ma*es a significant contri#ution to the (%imensional structure of nucleic aci%s.
• !romatic #ases a#sor# G light. – G spectroscopy can #e use% to pro#e structural transitions
in nucleic aci%s.
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1.1I) automerism
he #ases can e6ist in multiple tautomeric forms.
N
N N
N
X
H2N
H
O
N
N N
N
X
H2N
O
H
Aeto tautomer $J.K)
7nol tautomer LI.1K)
H
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iClic*er 8uestion%n the abo&e e'uilibrium why does the (eto
tautomer predominate o&er the enol tautomer?!. he stan%ar% free energy of the *eto tautomer is greater than
the stan%ar% free energy of the enol tautomer.
B. he stan%ar% free energy of the *eto tautomer is less than thestan%ar% free energy of the enol tautomer.
C. he O(' #on% is more polarize% than the N(' #on%.
D. he *eto tautomer is #etter a#le to form a atson(Cric*#asepair than is the enol tautomer.
7. Both B an% C.
N
N N
N
X
H2N
H
O
N
N N
N
X
H2N
OH
*eto enol
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1.11) he Phosphates
pAa = 2 pAa = @ pAa = 12
• Phosphoric aci% contains three ioniza#le o6ygens
• Phosphates polymerize #y formation of anhy%ri%e lin*ages
pyrophosphate
pAas of pyrophosphoric aci% = 1& 2.4& ?.1& F.4
•
Phosphates :oin to sugars #y formation of phosphoester lin*ages
$e.g.& 25& 5& or 45 O of ri#ose) Monoester pAas 1& ?Diester pAa 1
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1.12) hat you shoul% *no+ from sli%es 1.(1.11
E. 3tructures an% nomenclature to *no+ from memory
!. he #ases sho+n on sli%e 1.F& along +ith theirnucleosi%e an% nucleoti%e %eriaties& inclu%ing atomnum#ering& stereochemistry& an% ionization states.
B. Primary structure of "N! an% DN! stran%s& inclu%ing
correct stereochemistry an% ionization states& an% themeaning of stran% polarity. ou shoul% #e a#le torecognize the 5 an% 45 en%s.
EE. he implications of #ase aromaticity on
!. Nitrogen #asicityB. he potential for stac*ing
EEE. Aeto(enol tautomerism ( the relatie sta#ility of thetautomers an% the significance of this to #asepairing
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)ight *anded anti+parallel
double heli,
1.1) atson an% Cric* Mo%el
81 Base% on a#oe %ata& ho+ many
#asepairs are there per turn82 Base% on the a#oe %ata& +hat is thehelical pitch
*elical parameters-
atson& .D. an% Cric*& /.'.C. $14) Nature1@1& @@(@F&
hese are the alues that QC %e%uce% from /ran*lin5s fi#er%iffraction %ata. he actual alues for DN! in solution are a little%ifferent. /or e6ample& the aerage helical t+ist in solution ismore li*e 9R.
Helical rise 3$% &
Helical twist 3'(Diameter 2) &
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1.19) "osalin% /ran*lin5s S(ray %iffraction %ata
/ran*lin& ".'. an% Hosling&
".H. $14) Nature 1@1&@9I(@91
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.raggs /aw-: n 0 dsin .n = an integer λ = +aelength% = %istance #et+een ro+s of atoms2$θB) = angle of %iffraction that gies rise to constructie interferencehttpQQ+++.matter.org.u*Q%iffractionQgeometryQgeometryTofT%iffractionT#raggsTla+T1.htm
1.14) Gse of S(ray %iffraction to %etermine molecular %imensions
x
,eft picture constructie interference
"ight picture %estructie interference
En phase Out of phase
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1.1?) hy %o +e use S(rays to %eterminemolecular structure
0 3o the smallest %istance that can #e measure%#y pro#ing +ith electromagnetic ra%iation is halfthe +aelength.
0 Coalent #on%s are typically 1(2 U in length.0 3o in or%er to %etermine molecular structures at
atomic resolution& +e nee% S(rays.
S(ray crystallographers typically use copper S(ray tu#e $V = 1.491F U)
2dsinθ B
= nλ
d =nλ
2sinθ B
Smallest possiblevalue for d =λ
2
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1) ". /ran*lin DN! fi#er S(ray %iffraction %ata
2) /ran*lin5s %ensity measurements suggeste% a 2 or (stran%e% heli6.) Chargaff5s rules9) W H in *eto tautomeric forms
1.1@a) Enformation use% #y atson an% Cric* to #uil% their mo%el
*elical Pitch 0 !2 3
Cross suggests heli6 ( angle gies helical %iameter of 2I U
'elical "ise = .9 U/ran*lin& ".'. an% Hosling& ".H.$14) Nature 1@1& @9I(@91
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1.1@#) Emportance of *eto tautomers
3li%e 1.1I implies that *eto tautomers pre%ominate oer enoltautomers #y a ratio of J1III to 1. hat is the ei%ence0 Presuma#ly you coul% measure the eXuili#rium ratio #y NM" or some other
spectroscopic techniXue& #ut E +as una#le to fin% such measurements in theliterature.
0 atson an% Cric* relie% on Xuantum mechanical calculations to +hich they +erereferre% #y Caen%ish la#oratory colleague erry Donahue.
0 he *eto tautomeric state has since #een erifie% #y countless crystallographican% NM" stu%ies.
N
N N
N
X
H2N
H
O
N
N N
N
X
H2N
OH
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1.1@c) 7i%ence that *eto tautomers pre%ominate in io
Ef or H assume enol form then H #asepairs #ecome possi#le
0 his coul% result in mismatches %uring DN! replicationan% therefore in mutations.0 he fact that such mutations rarely occur erifies that the
enol tautomers rarely form in io.
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iClic*er 8uestionTo calculate the distance 4d5 associated with aspot in a fiber diffraction pattern which of the
following things do we NOT need to (now?
!. he %istance of the spot from the center of the %iffraction
pattern.
B. he %istance of the S(ray source from the specimen.
C. he %istance of the film from the specimen.
D. he +aelength of the ra%iation.
7. None of the a#oe.
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1 1) t C i * # i
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1.1) atson(Cric* #asepairsE +ant you to hae a soli% grasp of thesestructures.
hy Because an un%erstan%ing of thespecificity of #asepairing is critical to anun%erstan%ing of0 Nucleic aci% structure an% chemistry0 DN! replication0 DN! repair 0 ranscription
0 "N! processing0 ranslation
%n other words almost e&erything westudy in this course9
!s +e %iscuss topics li*e DN! replication&"N! processing& an% translation& +e5ll seethat the #asepair rules can sometimes #erela6e%. o un%erstan% ho+ they can #erela6e%& you first nee% to un%erstan% +hat%ictates this specificity in the first place.
*ence ne,t wee(s 'ui>999oet& oet& an% Pratt/igure 29(1
iCli * 8 ti
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iClic*er 8uestionWhy does the NA double heli, only
contain AT and @ basepairs? !. hese are the only pairs of #ases that hae
complementary arrays of '(#on%ing groups.
B. hese are the only pairs of #ases that fit into the %ou#leheli6 +ithout leaing any unsatisfie% '(#on% %onors oracceptors.
C. hese are the only #asepairs that e6hi#it perfect symmetry.
D. Both ! an% B.
7. !ll of the a#oe.
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1.2I) he pseu%o%ya% a6is
he glycosi%ic #on%s aresymmetrically arrange%aroun% the pseu%o%ya% a6is.3o the %ou#le heli6 smoothlyaccommo%ates !& !& HC&an% CH #asepairs
oet& oet& an% Pratt/igure 29(1
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1.21) hat accounts for the a#sence of non(atson(Cric* #asepairs in DN!
1 22) h t h l% * f li% 1 1 1 21
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1.22) hat you shoul% *no+ from sli%es 1.1(1.21E. he atsonQCric* mo%el
!. hat +as the significance of the follo+ing in formulating the
mo%el1. "osalin% /ran*lin5s %ata an% Bragg5s la+
2. Chargaff5s rules
. Ano+le%ge of the correct tautomeric states
B. hat are the most important features of the mo%elEE. atsonQCric* #asepairs
!. ou shoul% #e a#le to %ra+ the ! an% HC #asepairs +ithgood geometry7
B. hat is the meaning of the pseu%o%ya% a6isC. hy %oes the %ou#le heli6 only accommo%ate these t+o
#asepairs
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1 29) Protein Data Ban* $PDB)
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1.29) Protein Data Ban* $PDB)Molecular graphics programs $such as mol an% Pymol) %ra+ structures of proteins an%nucleic aci%s using %ata in the Protein Data Ban*> $httpQQ+++.rcs#.org).- 7ach structure e6ists as a PDB file& a list of all the atoms +ith their coor%inates.- !s of 12Q2FQ2I14& 119&4? structures ha% #een %eposite% in the PDB.
6,perimental ethod0 S(ray structures Y 1I2&1410 NM" structures Y 11&2190 7M structures Y 22
0 'y#ri% Y F0 Other Y 1
1 2 ) 7 l f B DN! % ! DN!
http://www.rcsb.org/http://www.rcsb.org/
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PDB ED $year) ype of heli6 $length) $resolution) 3eXuence
1BN! $1F1) B(form $12mer ) $1. U) CHCH!!CHCHB37 $2II) B(form $1?mer ) $1.? U) !C!C!!HHC!!D $2I12) !(form $19mer ) $2.4 U) CCCCHH!CCHHHH1Z/1 $2II4) !(form $1Imer ) $1.4 U) CCHHHCCCHH
he Dic*erson Do%ecamer> Y the first right(han%e% %ou#le heli6 to haeits structure %etermine% at near atomic resolution
1.24) 76amples of B(DN! an% !(DN!structures in the PDB
What does a pdb file loo( li(e? /ets go to the pdb to
find out7
1 2?) , t5 P l t l * t B
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1.2?) ,et5s use Pymol to loo* at B(DN! an% !(DN! structures
e5ll a%%ress the follo+ing Xuestions1) hat are the ma:or an% minor grooes2) hat %o +e see in the grooes) hat is the position of the helical a6is relatie to the
#asepairs
9) hat is the sugar puc*er4) hat %o +e mean #y #asepair tilt?) hat %o +e mean #y propeller t+ist@) hat are the ma:or %ifferences #et+een B(DN! an% !(
DN!
e5ll also practice rea%ing the seXuence of DN! #y loo*ing intothe grooes
http://www.boatdesign.net/forums/attachments/props/19848d1207043400-propellor-pedal-powered-catamaran-cimg0421.jpghttp://www.boatdesign.net/forums/attachments/props/19848d1207043400-propellor-pedal-powered-catamaran-cimg0421.jpg
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1;BC
1BC
'elicala6is
1.2@) hy are the grooes of %ifferent +i%ths
1 2F) Differences #et een ! an% B DN!
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1.2F) Differences #et+een ! an% B(DN!
%mportant differences
1) !(DN! has less rise than B(DN! %ue& in part& to the %ifferent sugar puc*ers2) he #ases are shifte% a+ay from helical a6is in !(DN!
a) "esults in caernous ma:or grooe an% shallo+ minor grooe
#) "esults in ? U hole
) Base pairs %ramatically tilte% in !(DN!
hese aerages mas* a lot of aria#ilitye.g.& in the Dic*erson Do%ecamer& helicalt+ist aries from 2?R to 9R& +hile helicalrise aries from 2.F U to 9.9 U.
hile these alues are #ase% on fi#er %iffractionstu%ies& solution stu%ies %emonstrate that hR for B(DN! in solution is -1I.4 #p& correspon%ing to ahelical t+ist of 9R an% a helical pitch of 4.@ U.
B-DNA A-DNADiameter 20 Å 26 ÅHelical rise 3.4 Å 2.9 Å
Helical twist 36 *̊ 31˚Helical repeat(h )̊ 10* bp 11.6 bpHelical pitch 34 Å* 34 ÅTilt 6 ˚ 20 ˚Propellortwist 11 ˚ 12˚!"arP!c#er C2’-endo C3’-endo
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1.2) 7ffect of sugar puc*er on #ac*#one e6tension
oet& oet& an% Pratt /igures 29(4 an% 29(@
Butane$Ne+man pro:ection)
[≅
14IR $close toanti)
[ ≅ FIR $close to g()
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iClic*er 8uestion
What is this basepair? !) !B) !C) CH
D) HC7) None of the a#oe
B37
N@ O? N9
O9 N? N@4(me
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iClic*er 8uestionWhat is this basepair?
!) !
B) !C) CHD) HC7) None of the a#oe
B37
NO2
NO2N2
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1.2) 25(O' group results in steric cro+%ing
Cro+%ing is +orse in C25 en%o sugar puc*er
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iClic*er 8uestionWhy does water stabli>e
.+NA relati&e to A+NA?
!) Only the B(DN! minor grooe has appropriate '(#on%inggroups for '(#on%ing to +ater.
B) he narro+ minor grooe of !(DN! e6clu%es +ater +hile the#roa%er minor grooe of B(DN! a%mits +ater.
C) Both ! an% B.
D) None of the a#oe.
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1.) he atson(Cric* Mo%el
atson& .D. an% Cric*& /.'.C. $14) Nature1@1& @@(@F&
1 9) B( s Z(DN!
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1.9) B( s. Z(DN!
B(DN!
B37
Z(DN!
2DCH
1 4) Z s B DN! #asepair
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1.4) Z( s. B(DN! #asepair
syn
anti
C5 en%o
C25 en%o
anti
C25 en%o
anti
D2DCH
.1BN!
stionshy %o #ul*y su#stituents $e.g.& Br) at F position of purine ring faor Z(DN!hy %oes high salt faor Z(DN!
%f you can answer 'uestion " on P81 you understand e&erything you
need to (now about D+NA9
1.?) hat you shoul% *no+ from sli%es 1.2(1.4
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1.?) hat you shoul% *no+ from sli%es 1.2 1.4E. !( s. B(form helices
!. hat is the meaning of each of the parameters on sli%e 1.2F
B. 'o+ %o the oerall shape of !( an% B(form helices %iffer an% ho+%o the %ifferent sugar puc*ers contri#ute to this
C. hat is the role of +ater in sta#ilizing B( relatie to !(DN!
D. hat is the role of the 25O' group in sta#ilizing !( relatie to B("N!
7. 'o+ is the atson(Cric* mo%el a hy#ri% #et+een !( an% B(DN!EE. Z(form helices
!. hy are Z(form helices restricte% to seXuences that alternate#et+een purine an% pyrimi%ine resi%ues
B. hat factors influence the sta#ility of Z(form helices
EEE. ou shoul% #e a#le to rea% the seXuence of a piece of DN! #yloo*ing into the grooes.