On the Translation of the Genetic Coce ; MARSHALL . NIRENBERG NaCionai Heart Irrm%&xde - National I. , Institute 5 of Health I ` ? , sis for a continuous dialogue between ceils and their descend- +e t.i ., t :I I.,. ants. Fossil records @bacteria:,about 3 billion years n&d I_! :' 1 . . ;;ave been reported @J$ BaghoorniSchopf); the first vertebrate Appeared approximately'500 million years ago; ani amphibians and mammals about 350 and 180 mi22ion years ago, respectively. ,Y The presence of bacteria 3 billion years ago m&-&&cate the presence of an operational code at that tixe# f k d lmost surely tfie code has functioned for more than 500 million years. The c; - >! remarkabie similarity in codeiwords used in bacterial, amphi- bian and mammalian replicative processes suggests that most, if not all, forms of life on this planet use almost the same genetic iangusge, and that this Panguage has been used, pos- sibly wi-;h few major changes, for at ieast 500 million years. : 1% _ i i ic Is by virtue of thi's language that izach generation Ls able co pass to the next generation a library of information whLch specifies in detail how to maIce the many kinds of protein catalystJthar: t'he ce22s wi.22 need for thefr deve2opment. cftrrdt-- M.t' now seems clear that all, or almost a22~form.s of life on this 7 lanet use virtualiy the same Panguage, vtly -.___ _ a number of "dialectsso have been found. ..I &a&-L describe thf."S The eluci Cation of the genetic code has been the subj/ecc. : i i of muc'n intensive work, particularly in the past four or <'five years, ar.d I woc.ld LL'ke to stress- . . . +-* C-at this w. rk ? ~~~Y&,Q,&&&~ .<,.f& e work with which I have,been associ- / &)\has been.' x- in a very real sense a colL.aborative project. ,i" * : G+-&&h--tki:s w-F~2&-~becorrte~~~~ as I proceed. : -_ . i.:. .A.' ,-,:..*a __--.. ' _ _,^ - *-. -- - $z&.++--q;;yyf~ r&c&,& briefly - as mual:: have be& , catalyses the synt'nesis of messenger- g thz DNA as a template. Only one strand of $&III DKA F. 4: < is copied by &C mA-polymerase; a~& the copying process is se- quentiall ,-J and -z%er~~T@sfgnaPs, whose exact nature is unkown, A@*s. G&&I. specify the beginzing and the end of.&'&-RNA t.r c * 2) ) ~s~hows. schematically%' ;he' T--q n process of protein. synthesis. In the DNA showil here, the dif- tc rerenc cross-hatchings represent various seg;nents of DNA, each (. i corresponding to a specific protein/or gro*cip 05 proteins. Xi- 1:"~ w. ,' Losories are shown,s-chemaG%calL& attached to the messenger~~~~A. 2: -4; ./_ - - ..c u&a c 05 collaborators at Bethesda.' -'.- i (,; ' ',"i_. ; _a wnc: ,e rsading, or translationibegins; as soon as one ri2ls- some moves down the - RNA, another becomes attached un- .,I I specific amino acids and recogniz parti- . Thus the GX&MWM&, 7 -,, -"----~gv .: iilustrates, H! 7 again diagraqtically but in more dzta,L. the codon recognition process as exemplified by that most inten- _-_ ts: the larger. XNA lies on the -pgtrs of tThe ri'bosome, can then bind5 ; i . . /yat one of two possible binding sites on the larger ribosomgsub- unit, a--particular amino-acid, (aa)-. One of these binding sites is for it3 tha peptidai s&NA, ~-,a 4he .d s+WA which is att'ached to the ._o growing (protein) ,poly&eptide molecule.; and the other/;for the ..- ..~ \ incoming amino-kid s(RNA. -%a~ hree enzymes ~.&he--two-~~-JUA!~ I-- L2.5 tI~-~e acti- vacior, energy are *requLred for the I transfer of the growing po'y.! c pe2zide chain to the next (incoming) amino-acid s?RXA complex. . _c Iv'1lcn this is Lecoxplished the srRNA required for the previous _c .r \ ,3 3 . .' * " amino acid is discarded, and a shift k-seme-by occurs so that the next codon (triplet of bases) on the x$RNA can be recognized -.. by a new s~fklL4. In this way the protein synthesis starts at a n,+s,7i ,.;.i.. `i:~+ t#, I; `2 given p 2ace (on the m$RNA), r&k+ti"groupings of ,thize* bases 4e- w&L& and with a given po2arity. : .` ; ,,'. -f---f ,- ,a f ? +~'*' ',y 6 ' #&p . i ' 4 In an actual living cell, even W bacterial'cel;, .j- `--+L), <" I_j +cci, , CT- Hb%chemical L&i- '- 3. processes a-333 simultaneouslyy~~~ all part of the cell metabolism, The synthesis of even a single i a\ .I ;Jrotein is quke an e2aborate process involving,'\jn-ter'-alia, tbi,e -I- , * ,,.&-..;fer of a long DNA message to an m@LNA molecule which .w-r it j ,, `, ' f, c:';-.aj: t-&L&y sufficient nucleotides (about 1,500) to code -me a, z ,A L..I Ii- ' &Sk amino-&ids - I, , cc I A :..: .:.'i 4 / f. p .- ?. :0X the-p-rote-kn ~p&k$~p+&Se --ch-a~~~,~ 'L.< Koreover. in an act-da!. ce22 these 2-1 any simple sequence, 1,500 nucleotides will not be arranged , I. .( ,I) (.r., . I. ref2eceing the fact that there is a whi.ch'.const-itute different proteins-*. Nonetheless p-#y /3-\ a ,-xt'variety of biochemical and genetic r;`- '.._ investigations, espec'%`aliy with bacteria and viruses, \\ 'many features of * protein syr;tlhesLSIt'incl~'~ngj,-~ particu2arly j3i;3ph LnformatLx Gout the code2tas been obtaicedy' .s' i The WOr-K T sha21 I#& cesc;-lb&g is, however, characterized Sy the use of much ,o simpler, Ln vkro systems, Ii1 wnere the essentially chemfcai fez-cures of some of the bask steps in the whoPe process are studied. Th? ,i:: success of these mathodsir~ and the -A concurrer-r ..? ,7 f, results Wwi*ch chose from in vivo experiments, w&s+~wY~.~JT-~-~~~~~ - aS%S== will,1 hopeJdemonstrate how a physio-chemical or mole- cular basis c2n be found fez t'nc 'UZST&C processes governing such fundamentally biological phenomena as cell metabolism and repli- cation. The basis for our earlier work on the DNA-MA code was the t. ) I;se of synthetic messagesi' (in place, tL~z is, of actual rnTFUA) .M which ,were randomly oriented sequences of the four code letters, t 'ne) G.(Fnine), the four bases 2 *,c j6 zacteristic ,r* 0;. the code could be . . ..tibase compositions I ' ,c"Y-t, ;r; , -bases idiaddaek worcs. ---.. l.lUS the problerr, up to /' ,;r two or three years ago was ilke :, . LhaC Oi an anagram: we knew .the letters comprising the codedwords but n&the order of the letters within each word. 'i =*-& 9 `. _* * ,'... I. established! in several 1aboratories;that -' - l&j,.:. 2'" sJ-~-"chlr. --eede%a synthetic m-RNA, in particular poiyuri- k,(\ `. ', : ! %&&& `./ dylic acid*% .~~synthe~ic RNA with entirely U basesJ3/to a suit- ,' c ._' a5Z.e mixcure of ribosomes, s+FUAgs, enzymes, ATP A) GTP and amino- acids I the ?o?iy-U WINS selectively bindr$henylalanine siIir;Ad f-Y \,.e :, the particular " sf-XXA associated wizh the incorporation of '4.: :/ '. _., 2 amino-acid ;2her.ylalanfne in protein),, tc t?~-~z ribosomes. '\ My co:- c .__-_ ._._. . league,PhiZ.Lp LeGerj and I then speculated Low small a message (G L :he FSA typz j wocld direct the binding of siXXA to the ribo- s axe . Experiment showed t'nat only three bases were needed, that i.sYjvery small molecules comprising only the triplet itself would dLrect the binding of the appropriate amino-acid sTR% to the ribosomes. This provided a,rather simple route towards the deter-ination of the sequence of letters in,the RNA coded words. Our main problem was to devise suitable techniques for syn- thesizing tri?iets. At the time we started our work with such triplets, methods had been reported for making some 20 or 25 of the 64 (=43) triplets which can be constructed from the four nucleotides Us C, ArG. These had been prepared by enzymatic breakdown of XXA, or by chemical synt'nesis, in the latter case using some of the very elegant techniques devised by Khorana and his associates. Two general techniques were developed in our laboratory, the first by Leder, Singer and Brlmacombe, a-rci the second by Xertoc 3ernfieI.d. The first employed polynuc:eotide phosphory- / x-f/L lzse ~$2.2 enzyme A&& % .'/ : &q! /& L;h$l;-igA' .JG- single nucieotides T v#.J,$>;;[~; .&/ &+&&$g&/ 1: 7- / to ditnucleotides to make ..I Fig. ? Zrimers , tetzamers, pentamers, etc. The second method em- ployed the enzyme pancreatic RXA-ase, which, although normally a breakdown or degradative enzyme9 will also catalyze an ex- change reaction between polynucleotides and car, be T;sed to make ",L - -? : bA Iplezs with ;JeLi-defined sequences. i3skTg t7z2 mthods of Khorana and these two enzymatic techniques, it was possible to synthesize amost all of the 64 trip ts. - * tp P I In connecCicn wL~h the use of N. small ~01ynuc~corLdc or "oligonuc&eotide" molec*ulss such as the trinucleotLdes, it LS important to-pain" i out LW that any given sequence of nucleo- tides can exis.c, when incorporated in actual miKXA in three chemicaily distinct fo-rms, depending on the location of the se- quence in the wkle messenger molecule. The chemical forms re- late to th2 three positions (a) as an internal codon (trinucleo- zide) 0;' as on.2 of the other of the terminal groups - so tailed 3i -termina 1 codon and 5' -terminal codon. -r'; This is ilfustrated in Pig. . Fig. All of 52 evidence //-.I 2; y : t _ ts+a&e suggests that the bioiogical char- ac~er~s,ics 02 eodon recognition may in some, perhaps in many, a, -1 :. : ir 1' , csuxts be influenced by the particular position of the codon in the rn.$,RNA (or equivalently in the DNA). Thus each of the 64 -triplets rsfercsd to above hay exist in three effectLvely dif- ferent structural forms. T=>Z significance of these "secondary" chemicai features ' 1s indicated by c* C? experiments/ in vitro,` ___._..l -._ . ..__ wit'? th2 GLigonucleotides > A ykL-,e he;ie;li :<.p* &;& 2 r ,.;JP/ with a methyl group attached to the 5' phosphate, and alsolwz The binding 02 -:he s'$3?!A to the ribosome is determked by tee::- :nic;ues ir: whit?. 2 Y2cii02ctive tracer -7 .-.a- .. is incar;oraced in the s,$.WA, SC, LndL t.-.Z rai- .Loactbvlty associated finally with the rLbosorAe zom?lex is & p,~Lsi;;"e c/f ---.k this binding. It is ic ,,y:h&t the telrr;l . . ..> / ;c-civity LA-- fiG-L;-c d2FAotes the eafectiveness al:." '- f" ,, ti. -II,,2 binding. , y *.b t' . ; <, .i-', ..- -- I , " (* . -,f I" 2 -7 7 be modified in such a manner. Certainly a substitution at the J 'i-terminus may be important because this could furnish a sig- nal which d r. s;Jec;rles the attachment and/or the detachment of the ribosome from h,`ne message, (m!RNA or subs:Ltute). Xecently Xtra and Surwitz, and also have shown that, in vitro at least, rn&+enge+lX..A contains a triphosphate attached to the terminal h;lyGTOXyl; and although it is not clear what physiological func- tion this zriphos?hate serves, it is highly plausible that it may in sor.2 way specify the initiation of reading the message. It cou'-' &ti aLso determine the first (three letter) word to be reai; ,' phase the reading, P and,; perhaps affect the susceptibility :G enzjrxs that could attack the te-rmini of the m---IA. internal codons may also be modified by these secondary chemical changes; the 2* hydroxyl or the base could be modi- fied and such c;?snges may be relevant to the punctuation of the T..j. -. ,_ * _ ._; 0 * ; : ;.: message. L==&so-cannot be excluded that the codon recognition process is In some instances affected by the ;jarticular neigh- bors of that cq> URGE on the message. CL;. .Jt I-t---sh&.d also.. b.e pointed. out that there could; * pxmeddy be a difference between internal initiation and termination (i.e.,. ., `.. IniclatLo;; or -:el%ination of polype?tLde sequence (protein) by a codon internzL,y iocz;zC -Lr, the message) and ?z:~z?zrl LzLtia- -1- Zion and t2XIii2~tiOn (th2 same process effect& by terminal co- t a 10 to coztaic the %nformstion for the assembly of more than one ?roteLn, ( or rnsre Char, o3e poiypeptlde c'nain of L --rotein). one starts Lo read (from the left in Fig. ) ehe codon for Fig. the terminal initiation, one then reads in the message until one reaches th2 word that says l'Sto$I,i and th-,en L&ze-wGG--be ,i $ 2.X UI; XT: GWT. mec%nism EG% start&$ the second message at an in- terval ?osltion. It seems quite piausible, although not known, that these -te;zrLnai and internal initiation and termination me- chsn L sxs co-u16 be different --possibly different codons. Anocher feature of codon recognition concerns t'ne degener- acy of the code, or the existence of synonyms, i.e. , different c codons which code the same amino 1" acid Ln the poly?eptide se- quence. XLth 1% e turn zow from these refinements arid detailed features or' the trl?lez- Sinding method to the actual results obtained bY this procedure. u the triplets have a well-defined se- rt J .E c*uence of nucleotides fthere are 64 possLbI.ee tri?Z.ets: a~& 5 we have synthesized 63 of these and detez;lz ed the aminobcids which they co* ._.,r "silent" 'Li i. %L'I,. t,. .-; a r? pew $ 8 .,i\i'. c. A-Y /i r , , mutation, that is [on one of the dower&, or gro?~ss __ r-:: .* ,. .` of synonym~cls code-words, td-,may be conver I, -d M&++ +~&+-&Kr&~-pos-it-~n to anotaer base without resulting in an amino 2, aciti replacement. Another obvious conclusion is chzt aminoA" acids wh',ch are very similar chemically, such as the dicarboxylic acids e' I aspartic acid and glutamic aci 9 >have closely related co,- dons. This may reflect the evolution of the code, but whether or not this isso, one consequence would certainly be that when an error in re?ilcation does occur, i;sually the first two bases are ,read Correctly ano - the third one incorrectly. And very of- ten the result of an error .in reading will be the substitution __ _ 4 .in a prozein~of a chemical+y related amino-+!acic$ . Thus the general picture of the code is that it is quite conservative-- in the sense that it usually minimizes error or the consequences of er- ror. The various .p.ttsrns bf synonym codons are summarized in FL ?? o (N-formyimethionine s'$UA shown here is the initiator). - In addition to the eoions for the specific amino/acids, &;-??ear to serve spec.lal functions ("punctuation" etc.). For zx- zqle, the recerr: work of Brenner, Garen and Zinder, knd of others, . 7. lnalca:iles chat UAA and UAG may indicate the end of a message - alt~Lough zhe precise mechanism for punctuation is unknown. of a message. i)ur recent s.tudies, and also those of Clark and Marker Ln England, have indicated that these codons - at i.east when in terminal positions - are recognized by formylmethi- onine and this may serve as a;1 Lni-zLator of protein synthesis. Soma p\=)ssl5ie s>eelal function codons are P2sted in Fig. . specify inktia-' LLon of message translation. And as I mentioned already, USG, AUG, CUG and to some extent GUG are recognized by A; also that UAA and UAG may serve as ter- ( -; , minators. It also appears likely that the words AG ~I,+&& ending'-*. 2, C, A or G may also serve as special function words; but'- i <>A ~.' these functions have not ~&f&c been found. The present situa- - :4.-f.. .i.- tion in this field is a most interesting one, In tZ-ia:,the neces- %%&, and 2-z sL-2.d soon be ?ossibPe to uSerstand more about the ;r.zzhanism sf thzsz special words and the role,-they pLay in pro- ,i tein spt:-.csis. &m-w+&L_o-l- turn--e a variation of the triplet- ";; :x&k binding method iW which--tkrow& further the coding mechan- iSXl. D. Xatfizld has recently prepared some radioactive trip- <-* ietsJ (in the ear 3.Ler experiments it was the s+.XW which con- tained the raGLo;ctive tracer),,"and has studied the binding of these trL-?iets to the ribosomes in the presence of the amino- acid szRXA. Fig. shows both the binding of the triplet and of the s:,RNA (here phenylalanine s,zR.iXA) to the ribosomc. Fig. UAW gl the presence of the appropriate triplet 2olynucleotide phenykaianlne G-RNA binds to the ribosome; in the absence of the s:tXNA very little triplet binds to the ribo- some. Because of this, in the presence of the s@NA both the triplet psiynuc' *eotide and the phenylalanine s$X.NA bind to the ribosome a~ ap>roximateiy the same rate. Thus the complex on the ribosose may well be a one-to-one association of triplet and T-his technique providek a very simple and quite sensitive method for detecting codon recognition by sjRhA w'nich is not acylated9' y.7 with amino'aci.ds. Thus some special function words may not be recongized by activating enzymes, s~XXA's, which are not ccykzed, and this method would provide a relatively sim;;;a route towards de:* ,ecting such recognition. We have also made investigations (in collaboration with 13. media contalnlng essentLally only a singLe type of STKXA, de- .' ^' r--c* ( . . j rived from Ejcoii fractions. We f&d that ,fyrosine-;jXNA re- _. . cognizes both UAC and ~-..,___ UA'd , which again exempliSLes 2-z C=U de- place. (There are two types of !Tyrosined,"' ; both types recognize UAC and UAU.) c-' u,milarlyl Vai,nelsfdmA recognizes both GUA and GUG (G=A degeneracy) but the GUG to a much lesser"extent than GUA. T'ne EIiicoli fraction u--`*. .- leucine-l-s-RX\ and leucice-2-s'%,RNA both :ecogcize the leucinz codons (UUA, LXG, CUU, CUC,'CUA, CUG). Recently, however, J. A. Carbon has re?orced that in mammalian 1Lver one species of i e-ic Ln e d y A 3.HA Treferentially recognizes L&G, and the other pre- ferentially recognizes SM. There are also types of leucine-s+,pA which recognize CUG, and others which recognize XX. The major variant of methionine f . . .stXNA which, as mentioned , previously > will accept a formy group recognizes UUG and CUG, -a.iJt a less prominer, me-Lkio~lne~siFQ?A recognizes A'JG preferen- ..!/ r;a1:y. LLkewLse there ' is a Aryp:o?han s1;t`Y.L TJ;YL ;, & recognizes UGG, CGG Lnd 1-3 a smaller extent AGG. T'r,e patterr, here is clear: a close relationship between U, C and A in the first place of 16 -- again the grou? U, C, or A but now in the third place of .?- -* ciL~ coding triplex. It should also ba poLnted o-ut p:omLnent % . .-. _" ;2~LX-LZlS-St*L\X b ir,ds to rlbosomes very weakly in response to the cogr; -L.;i on involves only two of the three nucfeotide basis in the x This work with pure fractions a'such as .a' alanine-si?A prepared from yeast, can afford some further insigFL-: into the mechanism sf codon recognition. This is especially 'i&in this case s;i;lce Soiley and l?.-iiS, collaborators have recently reported the sequence of bases in the alanine-s$,RNA. s i?ig. E&i show&" .a */ '_ Lhe'-'var-i;.at&on -o& binding of .alanine+stWA .\,.< i to ribosomes$&& con- I centration of the sIf possible interest. ,." I sequencet' 6, T,PU, 2 ($3 is 2n isomer of U)wm has been found in vir- tually every s$%A that has been examined. Another interesting seqiecce is the C, G, Gf7surrounded by two dihydrouridylic acids. A third is the TGC region (Hsinosine) right LE. the xiddle of .d.. Ce .stXNA molecule. .:.* These latter two regions of interest are 7 snown in mo-re detail in Fig. F?g. G&E, h " If u tri>iets CGG and IGC were really the sxRNA anti% codons, that is,the nucleotide groups which recognized the nu- b cleotidedtrip~;.~ ccs- ! :0-r alanine, recognition would be by Faral- lel" P&AA. ALLO = -c 2 '? 0` -2 2 ZT(JZ &J C and U; and the G woulf. th$n have to recognLze Jc ,,/ $ ,:+ s / I ,.. . ' .'.. q&!;.,,-, :_ {: f.. _ .., !,( ,:' . ::rrt: ..GC I U, C and A. YL, however, base pairing`M*accordLng to the 2 "I Watson-Crick hydrogen L onding, or antiTparalle1 scheme, C would d pair with G, G wi.th C and the inosine I in this position base-pair with on2 of U, C or A, but not G. This latter the pattern observed for the alanine code; and Crick has would is re- . . cently proposed a detailed mechanism which would permit hydrogen--'r" bonding between X and U or C or A. This mechanism, by which I recognizes U, C or A in the ,;i-..' * antitiodon - - codon pairing, termed the "wo';S1e",:by Crick, in- valves a movemmc, at the end position of the triplet, of either Z-ie smA or the rnw RNA on the ribosome. All the experi- mental resi;lts are, I believe, in accord with this type of re- g . . cognition 1,. - mecntinlsm. The table shows the based$equences in the Table s@A anti3odon and the corresponding basersequences in the mvX.NA codon. Thus Losine in an end position in smA ? I cdl&-~ C.?. L-P! ' can recognize by a-l-ternate base pairing U, C or A; a G in the -11. 'j ,. 6: ( ( ('J ,..iji 1' ,,J/&k :;;s end position of s&NA could'similarly recognize aL.-t~rnat:-el~ C or U,an-d A could recognize U, C or w and pb U could recognize ii ( ; ; . . . . (..$; : ' Q by alternate pairing A or G. . We would also predict on this mo- del t'nat a ri~;2-iymidylic acidi;mA woy.ld pair ;Iso A and G, c? erha...s j-he : .- ;-r -an ' ,..,~~ction with A wcjuLd be stronger than for a urLGylLc acid Ln s$CA) ; that a YU in s"zXXA might recognize al- ,^ +-- /-' i. J -.J_;, -/` ; J*i. f .-`, _, , ., , ,,& k.--A-a-c 2 -.i,i A, G 0;' g - a pattern that has beer, noticed rather Another possibility is #dihydrouridylic acid wcui-d not.,& kt .'I,._ i base 2ai.r ~&LL.~-L~ ",.~x~"~d-e~mplementary +-> ) so that the b.c.z it is also quite possible that a U or C Ln a terminal po- sitcon would not greatly inhibit the interaction. A metal grou? oc h 2l q-q&oxy~ ieoxyribose (sugar) mlg% also result In a weaker in:errction, and furthermore, by permitting a greater freedom of motion on the ribosome, such a modification might result in greater ambiguity, i.e. .J lower specificity of the coding. 3hes e * 7 resdcs wltn infrequently occurring (or "trace'? bases, and particul arly those with Inosine, i-strongly sug- gest c'hac s-WA may be modified enzymatically, after it is re- leased from the DNA template (where it is assembied in the cell). Since the level of sitrace" bases is quite high in an actual c~l.1~ iiz seems likely .:hat there exists a whole spectrum of interme- diates, s$,?.HA's in various 3tages of successive modification. The consequences of this are * easy to visualize. For ex- "i ampie, If an aienlne(A) Ln s@NA is de-sminalid and so converted in3 a~; Lnosine(Z)) t?.e A which would no-i?nally recognize the .4$ri- ct dyiic acid base in the message/would ncW be replaced by some- thing (the I) wh%ch can recognize U, C or A. si;yi;&+, intercon- versions would result from the deiamination of a C oz *:he conver- .`.,. 20 r SL0l-i 0; 2 t-J y. Tt is possible, although> -perhaps rather pre- ma tu-,`C -2 SpecLate, that this type of interconversion plays 2 es great deal of ET suggests ~ha,;t~ %,/is possible in actual .&.fJM+ ' fy ->Ae SpecificLty of ~_ . . . - ._ ,..._ I __ _ __ _ ._ codon recognitio+ a&-+++-- " ,`_ _:---. h--e -?%vP+rofound biological conseQ<*$, bian: @nosus laevus] liver, from guinea pig liver and from Eg -1_ 1-1 !T , coli. 2 -- dueoii argLiIineJs:MA does not recog- ---.-__". h>Llize AGG and recognizes CGG only very $7 r:htly, &?::eas 1.:~ both ..: -,-,`- -- 2 5 . . . .- '.?T, and mammalian for alanineAs,f,,., 2xTY& This contrasts with the activity for Eccoli -- ~- / alanine-s-t?XNA. --.q Ail all species testedlAM is recognized (*by dy- s ir, 2 c s jI-;O ..J fication of th5 s-LL~, there -% a cessation of proteL;: synthe- sis by the tat-terlai host. We do not understand She mechanism 1' rLr*i*g-ofQi s;.L 02 z'nis ,:,! but we think it 1ikeLy that,er,zyme pro- / .! * c,ced by T-2 infectron so modifies the leucisegs,.(RNA com-i)onenc as iz3 kterfere wLth the host protein synthesis, and it does this without preventing the protein synthesis by the phage. l-71 Inis is a very sr;btle way of subverting the metabolism of a cell 1 - how some features of the compiex mac'hinery for protein synthesis In ceils can be studied *by means of relatively much simpler systemsi' in vitro. Thus it has been established that the --- same sequer,ces of three nuc'leotide bases &e the same amino-,") acids throughout the whole range of organisms, from bacteria to mammalian llv~rs. And this universal code has been explored by molecular biozkemislry in vitro. -, . . . . . . . r. ,._l__l Xowever, we have seen that there aze secondary features, such as .;.Zie rklztive responses to d",fferent synonym codons, and 23 .~.y 2-2'1 ~;;:po;-~~i;-,-~ bioLogica roles; by selectively controllin,? 3 b,C -- .1 rCGLe ~- p-3,ein synthesis they may be an important factor -' ._ iri .ciie general process of cell differentiation. These are cer- tainly problems for the future. .(: it jlI . i..: ",Ju Finally, I we&-d-d -th++a~ that even, zn c c V 2 .L-* &LL 3, hC /e- its slm;?iest, the whole detailed process of c~Lk.g Ln ?roteir, synthesis - involving DNA-m@A-sTRXA-ri.bosGmes, L"..J activa- tion enzymes, AT?, etc. is far from fully understood. Even the basic underly'_;:A c;uestions - why, for example, does a triplet code of this sort exist, why should not phenylalanlne instead of ~~a;--L~~ CO--- ,,espond to GCU and GCC? Is there a basic che:::cal r -ZSCrSOZ 20~ thLs, or is it to some degree a matter of (ListorLeal) c ? CL-* c 2 7 . . ;t;y perssnal be lief is that there is an underlying mean- -e iA& for this a2 that it will be found.