Wobble as Clue?

On Wed, 10 Sep 2003 03:40:02 +0000 (UTC), [email hidden]

Quoted message said:

This tRNA is very curious, very curious indeed. I think it holds clues to the origin if we can
figure them out.

The wobble hypothesis suggests some things. The idea is that there is wiggle room in third position
codon base and first anticodon base. Allowed pairings include 3 position codon base / 1 position
anticodon base:
A / U,I
G / C,U
U /G,I
C / G,I

"All pairs of bases that can form hydrogen bonds are considered to be possible in the 3rd position
of the codon, except purine-purine base pairs, which would cause excessive distortion in the region
of the pairing." (General Genetics, Snyder, Freifelder, Hartl)

You show a I / A pairing and then quote no purine/purine pairings are possible? I and A are both
purines. The I/A pairing is allowed/used though it probably is more prone to error.

Quoted message said:

Now IF the purine-purine can't wobble, then it is probably because it is too fat to fit! But if it
is a problem with shape then it suggests the importance of shape/fit to the creation of the 3 base
codon and the genetic code. Isn't it likely that the code began as 3 bases because 3 bases fit (and
perhaps it needed a center base and 3 is the minimum to get a center position)

Also wobble in the 3rd position suggests a 3 codon base system that used to be 2. But why stop
there? Could that suggest the natural progression of 3 came from 2 came from 1? I think so. That
suggests that one (and because the center one is so key to determining the coding) base the center
base was the key to the origin of the code.

But how can 1 center base code? If A/U then it can. A, in center position = hydrophilic Amino acids
U, in center position = hydrophobic Amino acids.

What then about the other positions. As long as there is not a purine-purine bump, it would
probably fit fine.

Comment on wobble?

[snip]

(snipped)

Quoted message said:

The wobble hypothesis suggests some things. The idea is that there is wiggle room in third position
codon base and first anticodon base. Allowed pairings include 3 position codon base / 1 position
anticodon base:
A / U,I
G / C,U
U /G,I
C / G,I

"All pairs of bases that can form hydrogen bonds are considered to be possible in the 3rd position
of the codon, except purine-purine base pairs, which would cause excessive distortion in the region
of the pairing." (General Genetics, Snyder, Freifelder, Hartl)

You show a I / A pairing and then quote no purine/purine pairings are possible? I and A are both
purines. The I/A pairing is allowed/used though it probably is more prone to error.

I checked to make sure the quotes were as written. You've caught Mr. S, F, and H, in an error!

Any comment on the idea of coding starting with 3 bases because it best fit, and the center one
being key as it suggests either hydrophilic or hydrophobic aa's?

Any comment on T. H Jukes work in this area?

TH

On Fri, 12 Sep 2003 00:11:53 +0000 (UTC), [email hidden]

(TomHendricks474) said:

(snipped)

Quoted message said:

The wobble hypothesis suggests some things. The idea is that there is wiggle room in third
position codon base and first anticodon base. Allowed pairings include 3 position codon base / 1
position anticodon base:
A / U,I
G / C,U
U /G,I
C / G,I

"All pairs of bases that can form hydrogen bonds are considered to be possible in the 3rd position
of the codon, except purine-purine base pairs, which would cause excessive distortion in the
region of the pairing." (General Genetics, Snyder, Freifelder, Hartl)

You show a I / A pairing and then quote no purine/purine pairings are possible? I and A are both
purines. The I/A pairing is allowed/used though it probably is more prone to error.

I checked to make sure the quotes were as written. You've caught Mr. S, F, and H, in an error!

Any comment on the idea of coding starting with 3 bases because it best fit, and the center one
being key as it suggests either hydrophilic or hydrophobic aa's?

I would agree that probably this 3-base ratchet mechanism is based on the fit at the anti-codon
loop. I see this mechanism as evolving before there was any coding. So, for me, coding didn't
have anything to do with it initially anyway. I don't like your use of your 'thermal cycle' to
explain the evolution of the code. I think our views are at odds here. I see it evolving late in
the RNA world where cells have complex metabolisms and are making their C,U,G, and A as needed
using chemical pathways similiar as done now. I also don't like your emphasis on A and U. I don't
even understand how you think C or G in the 2nd position would be handled. I do agree that there
is a hydrophobic/hydrophilic split on the 2nd position but I see a purine(GA)/pyrimidine(CU)
split not an
A/U. I like that you seem to start with a very simple beginning which is the way I like to envision
the code beginning. The simplest possible start for coding requires 2 aminoacylation sites with
2 species of tRNA with rejection of some tRNAs during translation. I like to think even simpler
where it first began with only 1 aminoacylation site which means there was no coding initially
since there is nothing to select from. With 2 sites you can have your hydrophobic/hydrophilic
split. The question then is what tRNAs are being rejected during translation. I don't think the
Watson-Crick-wobble descriminator mechanism was in place at the beginning. I don't think you
are familiar with this mechanism which involves flipping out A1492,A1493, and G530 of the 16s
rRNA which must h-bond to the top edge of the codon-anticodon pairings. This has all been
discovered in the last few years. It can be blocked and when it is almost no tRNAs are
rejected. This mechanism came early but I doubt it was there at the beginning. I would
speculate that without this mechanism the only rejection of tRNAs is the size distortion in the
2nd postion from purine/purine or pyrimidine/pyrimidine pairings with the outside positions
being more tolerant of these size distortions. Eventually this should be known one way or the
other. There has to be something about the 2nd position that centers the
hydrophobic/hydrophilic split on it.

Quoted message said:

Any comment on T. H Jukes work in this area?

He was always full of ideas about the genetic code. If one idea didn't work he would come up with
another. It that way he seems a little like you Tom. I think he looked too much for explanations of
patterns though and did not look enough for initial simplicity. William L Hunt

Quoted message said:

TH

[snip] ...

Quoted message said:

WLH I do agree that there is a hydrophobic/hydrophilic split on the 2nd position but I see a
purine(GA)/pyrimidine(CU) split not an
A/U.

TH I 'm looking at this textbook chart, "Classification of 18 AA's according to the nucleotide
found at their second codon positions and known hydration potentials of their side chains." And it
is clear to me that second codon U (in many cases) is strongly hydrophobic with second codon a (in
most all cases) is strongly hydrophilic. I don't see how GC would work in this position. There are
G and C cases like GLY that are highly hydrophobic, but they don't seem to be as plentiful. And C
only ALA is hydrophobic at all.

I agree that 2nd position U codes the most hydrophobic and position A the most hydrophilic. I might
disagree on calling glycine a strongly hydrophobic. Usually it is considered hydrophilic but
sometimes neither hydrophobic or hydrophilic. Also (under C) proline and threonine are seen by some
measures as slightly hydrophobic and by others as slightly hydrophilic. Mostly, I see the initial
split as coming from having two aminoacylation sites (rather than current 20), one generally
favoring hydrophobic amino acids (UC 2nd position) and one hydrophilic (AG) but not specific to any
single amino acid. There need be but two species of tRNA. One with an A73 and a G35 and the other
than an A73 and using a U35. Position 73 would be the first aminoacylation site discriminator.
Today Serine and Tryptophan are the exceptions to this A73 rule. At the codon/anticodon, before any
Watson-Crick discrminator mechanism, the only tRNA rejection might be just a 2nd postion size
distortion pairing, purine:purine or pyrimidine:pyrimidine. So codons with 2nd postion U or C would
use tRNAs from aminoacylation site 1 (the generally hydrophobic) and codons with a 2nd position G
or A would use the tRNAs from site 2 (the hydrophilics).

... [snip] ...

Quoted message said:

WLH I would speculate that without this mechanism the only rejection of tRNAs is the size
distortion in the 2nd postion from purine/purine or pyrimidine/pyrimidine pairings with the outside
positions being more tolerant of these size distortions. TH I agree with the 2nd position. I agree
on size distortion. But I don't understand the pur/pur or pyr/pyr pairings idea. I think the stop
and start codons lead us to my A/U idea as more likely. Could you explain your idea further. I
think it may be the key difference between us.

I don't see the start and stop codons as more than later tweaks. The absence of any tRNA for a
given codon probably was enough to create a stop without any protein 'release factor'. These
protein 'release factors' had to come later. The question is why did these particular codons go
from coding an amino acid to becoming disconnected from any tRNA?

Quoted message said:

WLH Eventually this should be known one way or the other. There has to be something about the 2nd
position that centers the hydrophobic/hydrophilic split on it. TH Agreed.

Tom Hendricks >>

On Mon, 22 Sep 2003 15:46:44 +0000 (UTC), [email hidden]

(TomHendricks474) said:

<< Mostly, I see the initial split as coming from having two aminoacylation sites (rather than
current 20), one generally favoring hydrophobic amino acids (UC 2nd position) and one hydrophilic
(AG) but not specific to any single amino acid. There need be but two species of tRNA. One with an
A73 and a G35 and the other than an A73 and using a U35. Position 73 would be the first
aminoacylation site discriminator. Today Serine and Tryptophan are the exceptions to this A73 rule.
At the codon/anticodon, before any Watson-Crick discrminator mechanism, the only tRNA rejection
might be just a 2nd postion size distortion pairing, purine:purine or pyrimidine:pyrimidine. So
codons with 2nd postion U or C would use tRNAs from aminoacylation site 1 (the generally
hydrophobic) and codons with a 2nd position G or A would use the tRNAs from site 2 (the
hydrophilics).

Quoted message said:

Quoted message said:

I think this has a lot of merit. If the bonding to the AA (at the 73) was at first some type of
hydrogen bond, or some type of similarly thermally stable bond as those of the codon/anticodon
(you're middle positon 35)

If so this would set up two variants. A 2 hydrogen bonded U or a 3 hydrogen bonded G.

Then both ends would be thermally similar in that they would denature at the same time in a
heat cycle.

That means that there was hydrogen bonding to the AA before the present method. And that there was
a similar bonding at the other end - the anti codon, codon end.

The coding would not have to be exact. It would have to have the same number of hydrogen bonds to
work whether in one two or all 3 places.

Ex. UUU at one end and a UAU at the other would both have a total of 6 hydrogen bonds to their
paired partners. OR If both sides were GC then there would be 9 hydrogen bonds to their paired
partners. 6 vs. 9 would set up two variants.

I note that U (with A) with 2 h bonds is related to hydrophobic with smaller side chains (fewer h
bonds there if it is possible to bond there), and the reverse is true with G with 3 h bonds =
hydrophillic = larger side chains.

This is surely a stretch, but it has possibilities in that it would finally suggest a symbiosis
between AA connection to the RNA and anticodon/codon coding.

Comment? Tom

If there was some h-bonding of an AA to cognate tRNA stem then there would be no need for initial
use of aminoacylation sites and the machinery would be simpler. The problem seems to be that RNA
aptamers that bind AAs are not small. A quote from Robin Knight, "...know amino acid binding sites
are much larger than a base triplet, and there is no evidence for direct binding between amino
acids and trinucleotides in solution." William L Hunt

On Thu, 25 Sep 2003 18:42:05 +0000 (UTC), [email hidden]

(TomHendricks474) said:

<< If there was some h-bonding of an AA to cognate tRNA stem then there would be no need for
initial use of aminoacylation sites and the machinery would be simpler. The problem seems to be
that RNA aptamers that bind AAs are not small. A quote from Robin Knight, "...know amino acid
binding sites are much larger than a base triplet, and there is no evidence for direct binding
between amino acids and trinucleotides in solution." William L Hunt

Quoted message said:

Quoted message said:

This presents problems for sure. I would think the present system would have been selected as an
improvement over any direct bonding if it happened. As environmental heat lowered, replacement
systems would be selected over those that used the heat cycle only.

I want to pursue it a bit because it would answer so much if correct. Do you, or anyone reading
this, see anyway that either 1 - or more likely two bases from a proto- acceptor stem or acceptor
loop (perhaps the stem was a loop that has evolved to a stem, through history) could connect to
AA's in this specific way:

G or C with 3 h bonds could bond to a hydrophobic AA and A or U with 2 h bonds could bond to a
hydrophilic AA

Or, even better, a pair.

IF this was not the system, we have a real problem in that the alterantive is that the system
popped up fully formed. I can't see that happening.

Do you have any alternative ideas?

TH

First, I have never liked the idea of h-bonding of AAs to a tRNA as an initial step with a switch
to a covalent bond coming later. As I have said you are not the first to propose this but it seems
to me the normal method of evolution would be to just refine such an h-bonding method and not to
make a big change to a covalent bond method. You see everything about the genetic code starting
much earlier than I do. I see it coming much later in the RNA world where ATP would be a basic
component of the metabolism and the use ATP to make this adenine-to-aa covalent bond would not be
a problem.

I'm going to give you my view on how this genetic code machine (ribosome-tRNA-aminoacylation site)
first came about since it is always in the back of my mind when I discuss anything about the
genetic code with you. I see the first machinery as building uncoded polypeptides rather than
coded polypeptides. An uncoded polypeptide is just a short sequence of random amino acids. In the
absence of coding the machine can be very simple. It is just a tRNA with three ribosome binding
sites to hold the tRNA in place while reactions occur. One is a proto-amnoacylation site that
attaches any amino acide to the adenine on the tRNA stem using ATP as the energy source. There
would be no discrimination between amino acids or species of tRNA as there is today. The other two
sites form the proto-ribosome where two charged tRNAs stems are brought close together. The energy
of a phosphate bond in a polypeptide is the same as the covalent bond of tRNA-to-aa so the amino
acid should spontaneously move from one stem to attach to the other aa to form an elogated
polypeptide. No energy is required to do this. The mRNA is not needed initially and I don't see it
had anything to do with coding at the start anyway. If was a later tweak that gave a direction to
the elongation of the polypeptide. Today tRNAs come first to the A-site and then translocate to
the P-site to give a direction to the polypeptide elongation. They are left unbound as they do
this 10-20 angstrom translocation from A-site to P-site. I see the first use of mRNA as a weak
binding that keeps the tRNA from exiting completely and guides it as it makes this small
translocation movement. I see it as having nothing to do with coding initially. Initially it was
meant to bind with every tRNA and reject no tRNA. Any rejection of a tRNA would just slow the
machine down since there is no coding there is no advantage to rejection of any tRNA. I see this
as starting very simple so that is not a problem for me. Even though coding has nothing to do with
the initial function of this machinery it will evolve coding quite naturally and incrementally and
for me that is the appeal of this model. The problem is these short random polypeptides must be of
sufficient benefit to keep this simple machine maintained via normal darwinian selection. This
means within these simple organisms the ribozymes must be sorting out the beneficial polypeptides
and discarding the others. I presume these short polypeptides would in some cases act as cofactors
to increase the catalytic capability of the ribozymes. The question for me is whether this simple
machine would be maintained via selection while building only short random sequence polypeptides.
I think it might be. William L Hunt

... [snip] ...

Quoted message said:

WLH I see it (the genetic code) coming much later in the RNA world where ATP would be a basic
component of the metabolism and the use ATP to make this adenine-to-aa covalent bond would not be a
problem. TH But this brings up the question, "what came before?' I don't see any reason why all
this would be coming together even at your later time period.

Just to be clear, I see these RNA world organisms as having hundred of genes if not more, that
transcribe into hundreds of different ribozymes that drive the metabolism and replication. Among
these ribozymes surely there is the tRNA, a tRNA binder, an an amino acid binding aptamer. It is
basic darwinian selection for these organisms. I don't know what the tRNA is being used for but it
is surely a basic component of these cells. So all the components for a machine to build small
random polypeptides are present, being used for something in the cells metabolism that maintains
them via darwinian selection. How we get to this state and "what came before" is another
discussion. Your model explains more how we get here and "what came before". I'm just starting at
this point where I see the machine for building short random polypeptides starting. You understand
that from the beginning (1st life) there was the thermal/hydration cycle that built these sort of
short random polypeptides. I presume these became incorporated into the metabolism of these earlier
organisms. The only need for an internal machine to build short random polypeptides at a cost in
energy comes when they become depleted in the "soup" and can't found for "free". I expect this
depletion of naturally formed polypeptides doesn't occur until later when these RNA world organisms
have greatly proliferated. William L Hunt ... [much sniped] ...

On Tue, 30 Sep 2003 18:38:08 +0000 (UTC), [email hidden]

(TomHendricks474) said:

<< You understand that from the beginning (1st life) there was the thermal/hydration cycle that
built these sort of short random polypeptides. I presume these became incorporated into the
metabolism of these earlier organisms. The only need for an internal machine to build short random
polypeptides at a cost in energy comes when they become depleted in the "soup" and can't found for
"free". I expect this depletion of naturally formed polypeptides doesn't occur until later when
these RNA world organisms have greatly proliferated. William L Hunt ...

TH I see where we are 'miles' apart in our timelines.

You mentioned in one post that others had thought of the proto tRNA being bonded directly to the
AA. Who were you thinking of? And what in the proto tRNA bonded to what in the AA in these people's
opinions? And if you will please make your chem explanations as simple as can be.

"Stereochemical" explanations for the genetic code involve some direct physiochemical interaction of
lesser or greater degree with an amino acid and a proto-tRNA (or mRNA). I see no need for a
"stereochemical" explanation but others are still looking evidence of this connection. You seem to
be looking at what these possible connections might be. Various proposals have been: AA to codon
(Woese 1969) AA to anti-codon (Wong 1988) AA to codon/anti-codon complex (Shimzu 1982) AA to stem
(Hopfield 1978)

With the discovery of ribozymes (Cech 1986) and later understanding that these ribozyme-binding-
amino acid "aptamers" were not small made these earlier proposals seem more difficult since they
imagined small binding sequences. Currently there is lab work in "evolving" these aptamers and
seeing what they look like. They are somewhat biased for purines (A and G). The most worked on and
most interesting are aptamers for arginine. These rna ribozyme aptamers binding the amino acid
arginine are in a range of about 30 to 60 bases in length. When all 3-base sequences in the
aptamers are compared to the actual codons for arginine, there is a very a high percentage that are
exactly the same as these arginine codons. This is way above anything expected by chance and
somewhat puzzling. When more work is done with other amino acid aptamers we should see if there is
some general pattern to this. Right now it appears the arginine aptamer may just be an anomaly. Of
course, if there was a general pattern seen with other aptamers also, it would beg for some
"stereochemical" explanation. William L Hunt

... [snip] ...