Some Foo-taotes on Protein Synthesis k Fate for the I?%+. Tic Club u_- __I. - by - F.H.C. Crick and S, "urenc2~ y_y_ -.- 7T ,;:.?oCc. Unit Cavendish Laboratory, Cailibridge, Engl32-d. Suinmary In this note xe -,?oint out: (1) that soluble RNA may have CL DIU-like secondary structure in order to cover up the if;;L&bptox seyuence[s xhen no amino xid ia ztti;Eched 50 the RNA 0 (2) that 2.t some stage the amino acid mz.y ?J~Z attached to the NT of the pse;':do-uri-1in.z of soluble H-A, (3) that taoT?,erticles ol c?;ifTlereu!t size 2nd shape can be constructed :rom identie.:l protlzin subunits if combixd ,rdith liNA ;-no? ,ZCT Aes of dil'ferznt length. In particular both the 50 S AM th-: 30 S ribosomal >artizlcs may be incolilpletc ?arts of :& splserical shell, -l- . The object of this brief note is to add a few details to some of our ideas on pro-kin synthesis. Soluble R8X The first idea that needs some revision is the "adaptor hypothesis". This originally sprang fro!yl the difficulty of conceiving an RKA tei>l?late with 20 specific cavities, one 'for each ai;lino acid. It was SU&;,_Lu c^rracted -ti?al; 20 specific "adaptor z.olec-klesvl would be req_/,ired to position the amino acids, and that each amine) :;cid would 17~; joined to its own special adaptor 'b;; a special enzyme. The nature of these adaptors was initially left opcn7 'out it YJL;;j later pointed out thzt the obvious choic;: ivzs 2, s;ilall polynucleotide which wo-dd combine 9 `0;~ bz,se=--paiyiLqFr; 9 with the groper pcLrts of the b s.:.se Sq~LLxlc~ 011 .-. tL;c P't~;Ilpl;,:,-t(Ly' FLQ\* '-?!11c subscquenv discovery of the 20 sp,ccific activating enzymes, and thsir role in transferring the ;XiliIlG acids specifically: to molGculcs of soluble RPJa at first supported these i~~~,s. Further ~;oy,'r;~ howev82r, has uncovered some difficul-ties. They arc: (1) The amino acids becoin atta,chcd to the terminal ribose of %hz soluble R?JH, yi>t the soqucncc of baszs at that end ap;,zars -la `~3~3 t-h.2 sa:1c for all (or almo,st all) LloleCLles Of soluble R&L 1 naiilelg aqdeyclirlc Zy/t o g ine (Cytosine). One might have expected that this pzrt of ths _ sequence mas s?ecific for each XiIino acid o (2) The soluble R%;L has a highdr molecular weight than T:jas expected.. Estimates vary from 50 to 130 bases, with 80 to 90 as the ;Lore likely values;. One might have expected any number grLz:ter than 2 but 1~3s~ than (9.~) 1-O. V/e can think of reasons for explaining th2 ne(;d for the common ter;i,inal sequence - tc fit on to a standard 2myne ', to prcvidc a link to bridge a distance 9 et-r=., - but as they arc all ra,l;;r oi:vious and as thz-r~z is nothing much to choosz bet-lJt?ex the>? ;;~e shall no-~ discus;; the;~ further here. On reflecting on the proble:J of ihc dL LJLA. r- ;x;-3:'r~n-tly excessive size of the soll;.ble RNA 9 ~CILVCV zr 7 it bocc:.z apparent that a rather i_nportant requl ' ruxnt httd Secn emit tad. It would -2- . delay protein synthesis unduly if an c;ilpL,y adaptor (Le, without its amino acid) could fit onto the teX1plate, and there was nothing in the original theory to prevent this. This difficulty can be got round in a nwiiber of wayss The obvious thing to do is to cover up the specific bas sequen@e 9 meded for. base pairing with the te17nplate 7 until the afi1in.o acid is attached. One rather neat 'iday of doing this suggests itself0 This is to make the soluble RNA fold back on itself to form a DnTii-like structure. (we know froin studies of '"Poly A plus Poly U?' that a polynucleotide with .& ribose backbone can taiL:e up a configuration like DNA.) 0 This requirezzent could well :account for the length of the RNA being greater than expected* The idea that soluble RNA may have sozne secondary structure is not new0 Dr. Paul Berg thought at one time that he had so;'i?e evidence for it (and 5309 more recently, hzve others) and it had bcen suggested to us in discussion by DY" Boman and by Dr, Bach. The only novelty in our ides; is that it suggests a reason for such a -- secondary structure9 if it exists@ The idea, then, is -that without the a;i?ino acid the molecules of soluble REX fold on thenselves so -that their "Adaptor se~uence'~ is covered. At sane time after the amino acid i): ?.ttsched this sequence is uncovered9 and can then base- paid with the te;xglate RRN;L of the ribosomes, Is there any evidence that soluble RnTA has secondary structure? This is too recent and co;ilplex a subject 'CO discuss hz-ret but we day note that the base coj:iposition of soluble RXA, deterr;ail?ed by three laboratories (Hendrick Ssrith, David Dunn and Jii;l Offengand 9 wl;if.th their various collsborators - personal communications) have showh that the base ratios tend towards the DN'.A rule ; that is -.- a .&:. U plus pseudo-U and G =A c. . (Incidentally if soluble RKA rese;:lbles 0%~ it raay act as a. te;zplate for its own replication) 0 There is one difficulty about this idea which is iriaginary. This is: how does the activating enzyme recognise the correct ijlolecules of S-RNA if the adaptor -3- se$ucnce is covered? The answer is that a protein can in principle rrcognisz: a pair of b:asesl by making contact in the deep groove of the Dllu'A-like structure, azhd so can distinguish one double-helix fro:;?. another7 with a different b 2:s': sc~uenc,~ 9 without neceh into a 50 S and a 30 S particle. Why should there be two unequal particles, and how are they related? Eobodg secins to have ;~roduced a very good reason for the two parts of -:hc 70 S particle, except tile idea that the 50 is a box and the 30 S is its lid, and that the two cone apart LlOLlCilt Z,1" il XT ri 1 :frc,i:, tic:; t 3 tii;;e 9 to let finished protein mokecules c stage . AS t0 th2 x~-mcr oi con?. kruction it ~~oubd not be surprising i% there `;J-;,s i^jy~z F;lotc?i;ri for t& 50 S particle and one for the 30 S, but recent !`;IGY~"; (The Harvmd Biology group - personal cormunicatior2) su~~sts that the protein of these two particles ;> h are rather sirrilz. I-i-i0 7; then do they corm to have such a diffe_rer,t size end shape'? Rather eur?risingly 1 a simple answer is possible, It is only mcessary to assume that the particle cannot easily be produced froz; p::oteir: sub-units alone, but needs RPL to stabilise the ary:~qgmc nt ~ This is in any case true for the rods of TIW* Sup-,os~s for the purposes of explanation, that Szf ore mse:I_:bly, the .RNA was cut exactly in half ~pcle ztight reasS3nab1.y mpect -kc Cind that h,::Cspheres would be produced instead c#f spheres e 011 this vicwT then, exh oi the two portions of a ribosome consists of part of 2 .-v-w spherical shell; the size of each would bo dc-iierrincd In;? Ihe size of the RNBa9 so that the protein/IUU ratio Tzould bc the sa;;le for both parts, as it found -do be the case 0 It is m-t osscntial for the protein sub-units of the two parts to l-12 idc~tlc& but one would expect ther_1 to be si&iikar. . Fig. 3 but this :v,,>!r:>r' 'iS: di_Zfictixlt to see why the two parts should not be equzlo .`-zher alternx,tivcs are : Fig. (a) (b) ;hese hz:~\i $Gz t'~-\-~;~.;~,g; that the contact between the two parts is ;_ZXZ, _z* fro;il t'?at between the sub-units 'of each part. Curiously enmx,-j'h rec,znt electron Ynicrogrqhs (Hugh Huxley - personal ~oixxn:l.cation) C!~O not look totally unlike Fig. 4(a) o It is -jn AilJT C;iL (3 ';YlGWE th&-t the smaller particles (30 S) is more asyXK!etri.~::~L -thzn the big,qer one (50 S). -7- 'This sort of idea would explain very dell why the m weight of the RNL has? apparently, two fixed sizes (lea aside th$z difficult question of the RNA breaking down into further sub-units) and not, as one Ught expect, a ~o~t~~~~~~ range of sizes. It 1-aises c0nsiderGbl.e difficulties when c0E.e~ to consider the Rl'iLA as z specific teiIlplate B ne is naturally inclined to the idez that only & of the rib mal RNA azt s a s a -te7iq+te 9 but further than that we shall not venture at thz zloilzent. . 4