Problem Set 4 on Central Dogma a b . In cells,

ChemicalBiology03 Fall2009 Goutte

Problem Set 4 on Central Dogma

(dueMondayOct.19th,inclassorby5:00pminProf.Goutte'soffice)

1. DefinethefollowingtermsastheyareusedinregularEnglishlanguageANDgivethespecial meaningeachtermhaswhenappliedtothelivingcell.

a. Replication: The action or process of reproducing or duplicating. In cells, the process of duplicating cellular material, such as DNA, to make two identical copies to pass on to daughter cells.

b. c. transcription:a:an arrangement of a musical composition for some

instrument or voice other than the original b : a recording (as on magnetic tape) made especially for use in radio broadcasting. In cells, the process of constructing a messenger RNA molecule using a DNA molecule as a template with resulting transfer of genetic information to the messenger RNA. d. translation:a : a rendering from one language into another. In cells, the process of forming a protein molecule at a ribosomal site of protein synthesis from information contained in messenger RNA. e. splicing:a : to unite (as two ropes) by interweaving the strands b : to unite (as lengths of magnetic tape) by overlapping and securing together two ends. In cells, the cellular process by which intervening sequences (introns) are removed from mRNA transcripts, and remaining exons are rejoined together.

2. Nowforsomenumbers...

a. HowmanydifferenttypesofnucleotidesarefoundinRNA?3 b. Howmanydifferenttypesofaminoacidsarefoundinproteins?20 c. Howmanynucleotidesareneededtocodeforasingleaminoacid?3 d. Howmanydifferentcodonsarethereinthegeneticcode?64 e. Howmanycodonscodeforaminoacids?61

3. Myoglobinis153aminoacidslong. a. Whatwouldbetheminimumnumberofnucleotidesrequiredtoencodemyoglobin?

(153x3)+3=462

b. WhymightthesizeofthemyoglobinprimaryRNAtranscriptbelargerthanyouranswer

above?Becausetheprimarytranscripthaslargeintronswhichare removedbeforethemRNAisusedfortranslationofmyoglobin.

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4. Usethegeneticcodetoidentifywhichofthefollowingnucleotidesequenceswouldcodefor thepolypeptidesequencearginine-glycine-aspartate: a. 5'-AGAGGAGAU-3' b. 5'-ACACCCACU-3' c. 5'-GGGAAAUUU-3' d. 5'-CGGGGUGAC-3'

SequencesaanddbothcodeforthepeptideArg-Gly-Asp.Becausethegeneticcodeis redundant,differentnucleotidesequencescanencodethesameaminoacidsequence.

5. Explainwhyifyouaregivenaproteinsequence,youcannotpredicttheexactRNAsequence

thatwasusedbythecelltogeneratethatsequence.

Becausethereismorethanonecodonthatcanspecifymostaminoacids,therefore youneverknowwhichonewasusedforaparticularprotein,unlessyoucanlookat theDNAorRNAsequence.

6. Considerthehypotheticalproteinrepresentedbysingleletteraminoaciddesignations:

A-M-H-E-R-S-T. a. ProposeoneRNAsequencethatcouldencodethissequence(don'tworryaboutstart

codonsforthisexercise).

LotsofPossibilitieshere!

b. CanyoufigureoutmathematicallyhowmanydifferentRNAsequencescouldcodefor

thisminiprotein? I'veindicatedthenumberofpossiblecodonsforeachaminoacid:

A(2),M(1),H(2),E(2),R(6),S(6),T(4).

InordertofigureoutthenumberofpossibledifferentmRNAs,yousimplymultiplythesenumbers (2x1x2x2x6x6x4),andyouget1152possibleRNAsequencesthatwouldcodefor"AMHERST"...now, justwhenyouarefeelingprettygoodaboutyouroddsofhavingfoundacorrectRNAsequenceto encodeAMHERST,youshouldconsiderthefollowing:howmanydifferentRNAsequencesare possibleforthislength(21nucleotide-long)RNA??(tofigurethisout:you'vegot4possible nucleotidesateachof21spots,sothat's421)Nowyou'llseethatifyouguessedrandomlyyou`dhave lessthanoneinabillionchanceofgettingitright!

7. Hereisthebeginningofaprotein-encodinggenesequence.

5'-ATGAAGTTTGGCACTTAA-3' 3'-TACTTCAAACCGTGAATT-5'

a. GivetheRNAtranscriptthatwouldbetranscribedoffofthebottomstrand:

5'-AUGAAGUUUGGCACUUAA-3'

b. TranslatethisRNAsequenceintoaproteinsequence.

MET-LYS-PHE-GLY-THR

c. GivetheRNAtranscriptthatwouldbetranscribedoffofthetopstrand:

5'-UUAAGUGCCAAACUUCAU-3'

d. TranslatethisRNAsequenceintoaproteinsequence.

ThisonedoesnothaveastartingMET,sotechnicallyitwouldnotgettranslated;ifit did,thesequencewouldhavebeen:LEU-SER-ALA-LYS-LEU-HIS

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e. Let'sassumethatthebottomstrandisthestrandthatisusedasatemplatestrand whenthisgenegetstranscribed.Whatwouldbetheeffectonthefinalprotein productifamutationcausedthefollowingsinglebase-pairinsertion:

5'-ATGAAGATTTGGCACTTAA-3' 3'-TACTTCTAAACCGTGAATT-5'

becausethereadingframeoftheproteinischanged,allcodonsafterthelysineare different,henceyougetacompletelydifferentprotein:MET-LYS-TRP-HIS-LEU

f. Whatwouldbetheeffectonthefinalproteinproductifamutationcausedthe followingsinglebase-pairsubstitution:

5'-ATGAAGTTCGGCACTTAA-3' 3'-TACTTCAAGCCGTGAATT-5'

nochange:MET-LYS-PHE-GLY-THR

g. Whatwouldbetheeffectonthefinalproteinproductifamutationcausedthe followingsinglebase-pairsubstitution:

5'-ATGAAGTTTCGCACTTAA-3' 3'-TACTTCAAAGCGTGAATT-5'

aproteinwithoneaminoacidchange:anargininereplacesaglycine;becauseatinya.a. isbeingreplacedbyalargechargeda.a.thischangeislikelytoaffecttheoverall3-D structureandhencethestability/functionoftheprotein: MET-LYS-PHE-ARG-THR

h. Whatwouldbetheeffectonthefinalproteinproductifamutationcausedthe followingsinglebase-pairsubstitution:

5'-ATGTAGTTTGGCACTTAA-3' 3'-TACATCAAACCGTGAATT-5'

thismutationchangesthesecondcodonintoastopcodonwhichhastheeffectof terminatingtranslation,thusnoproteinwouldbesynthesizedonthismessage (unlesstherehappenstobeanotherAUGcodonavailablefartheralong,andinthis casetheoutcomedependsonhowfarawayitis,andwhetherornotitisinthesame readingframeastheoriginalstartcodon).

i. Whichoftheabovemutation(s)wouldyouexpecttobethemostsevereintermsof theoveralleffectonthepersoncarryingsuchamutation?Explain.

(e)and(h)wouldbethemostsevere.Thereisaslightchancethatin(h)anotherin- framestartcodonexists,andthustheoriginalproteinmighthaveachanceofbeing madewithsomemissinga.a.atthebeginnings,butthisisnotthemostlikely scenario.(g)isonlyonea.a.differentfromtheoriginal,howeverifthishappenedto becriticalforproteinfolding,itispossiblethatthismutationwouldbejustasbadas (e)or(h).(f)wouldhaveabsolutelynoeffectonthepersoncarryingthismutation.

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