In a message dated 9/18/99 12:16:24 AM Mountain Daylight Time,
MikeBGene@aol.com writes:
> >In any event, the answer is self-organization: molecular structures,
> >using the physiochemical laws, are able to form larger, more complex,
> >even organized structures on their own.
>
> I must say that you have a far greater sense of certainty about
> these things than I do.
>
It's hard not to after reading the amount of research I have. This work has
been going on for four decades and at one time or another as many as 25
people world-wide have been investigating this one individual model. Even
many of its critics agree that this model has come the closest of all
available models to explaining the origin of life, and a significant number
are willing to accept that thermal protocells could be or even partially are
living structures in their own right.
>
> Self-organization may be the answer (and
> I suppose must be the answer if we exclude some form of
> intervention by an intelligent agent), but I can't say that it *is*
> the answer (although I think it does have a great deal of theological
> appeal).
>
At least as far as abiogenecists are concerned, I am aware of no one who
rejects self-organization as the basic foundation for the origin of life.
Virtually every scenario (that I know of) that has been proposed -- whether
it be based on protein first, genes first, hypercycle, what have you -- uses
self-organization as a working assumption, based on the results of
experiments. In many ways, self-organization forms the basis of modern
chemistry and biology. Why does a mixture of hydrogen and oxygen gas always
form water when ignited and not hydrogen peroxide or any of a huge number of
possible combinations? Unless you want to invoke "intelligent intervention"
(in which case I would want to see your evidence for this) the classical
answer is that the physiochemical structures and functions of hydrogen and
oxygen provide the information needed to force them to exclusively form water
every time. Why does a specific sequence of amino acids always fold into the
same three-dimensional shape? Experimental evidence has told us that it is
because the R-groups of the amino acids direct the folding of the polypeptide
chain. The amino acids associate to form secondary structures, these
associate to form functional domains and these in turn associate to form the
overall three-dimensional structure. The experimental evidence supports the
same basic classical answer that is used to explain the formation of water
from hydrogen and oxygen gas.
In both examples the classical answer is self-organization based on specific
structures and functions mediated by the physiochemical laws. The only
difference is that in the former case we know precisely how the
self-organization of water works, whereas in the latter case we still do not
understand how and why proteins fold the way they do. Yet self-organization
is still the basic explanation for virtually any physiochemical process, even
chaotic ones. If self-organization is fundamental to any physiochemical
process, then if abiogenesis is a physiochemical process self-organization
must be fundamental to it as well.
As for "theological appeal", I cannot speak for anyone but myself. As a
theist I would in fact tend to reject any concept that claimed that God did
not create the universe. As a scientist, I am required to accept what
experimental evidence and theory explain about the natural universe. Since
science indicates that self-organization forms a fundamental basis for the
natural universe, my theology is that God created self-organization and used
it as a tool for creating and maintaining the universe. As such, if I were
not a scientist, my theism would in fact tend to have me reject
self-organization, so in my case at least self-organization holds no
"theological appeal" for me; if anything I find it theologically
contradictory.
>
> >For example, heating amino acids in the absence of water creates
> >polypeptiditic macromolecules with non-random sequences that
> >are catalytically active; these are called thermal proteins (many
> >people still use the old term proteinoid). These thermal proteins
> >are then able to form cellular structures upon rehydration that can
> >convert sunlight into ATP, create polynucleotides using thermal
> >proteins as templates, and create polypeptides from polynucleotides.
> >They can also perform other metabolic reactions, they can grow
> >and they can reproduce.
>
> Very interesting. Since these cells grow and reproduce, I would
> assume someone has a culture that has been frozen (a common
> procedure for microbiologists). Who can I write to in order to
> get my hands on these "cellular structures?"
>
This is one of those good ideas that should probably be done but hasn't been
done yet, though in this case I'm not sure if it would provide any meaningful
results. The ability to survive freeze-thaw is not an accepted cellular
trait; in fact, if anything, cells are well-known for NOT surviving
freeze-thaw, unless special precaustions are taken. Proteins are more likely
to survive, but still many do not unless they are lyophilized first or
protected by salt, sucrose or detergeant buffers. As such, I would predict
that protocells made of thermal protein would be distupted by freezing,
though they may reform upon thawing. On top of that, thermal protocells are
structurally like modern cells -- basically hollow spheres with a bilayer
boundary. As such, like modern cells, thermal proteins would also be
vulnerable to ice formation inside them; since ice expands as it forms, ice
crystals tend to disrupt any cellular structures.
There are really only two reasons why researchers freeze cultures away. The
first is if getting more fresh cells is difficult or impossible. Otherwise
researchers prefer to simply go back to their original source for more cells
or instead raise cells known to be very hardy, like fibroblasts or various
strains of cancerous cells or bacteria. The second is to preserve any cell
lines that have interesting properties.
In the case of thermal protocells, since it is very easy to make more
protocells and relatively easy to control the properties of those protocells,
it hasn't been worth the trouble or effort to try to freeze protocells away
for future use; instead it has simply been easier to make more as needed.
(It would, however, be an interesting project.)
>
> Furthermore, how
> would I go about making these cells? Can I just weigh out a few
> grams of randomly chosen amino acids, put them on a heating
> plate for a few minutes, add water, and observe? Or is the
> procedure more specific than this?
>
Yes, but not too much more. The classical method goes like this: mix one
gram of aspartate and one gram of glutamate with one gram of an equimolar
mixture of all remaining amino acids except asparagine and glutamine (the
heating process tends to convert them back to aspartate and glutamate). Heat
them at 180 C for one to six hours. Meanwhile prepare a 1% NaCl solution.
Bring it to a boil, then carefully pour it over the heated amino acid
mixture. Dissolve the mixture, bring it to a boil for a minute or two, then
allow it to cool. The protocells will form from the dissolved thermal
proteins as the liquid cools. You will need a microscope with a
high-powered, oil immersion objective to see them, though.
As I said, this is the classical method. You get the same result by using
various proportions (say, 2:2:3), you can substitute lysine for the
dicarboxylic amino acids, you can use just aspartate or glutamate instead of
both, you can use an equimolar mixture of all 18 amino acids (again leaving
out asparagine and glutamine) with no excess of either dicarboxylic amino
acids or lysine (for example using a gram of each amino acid), and there is a
possibility that glycine alone (in the absence of aspartate, glutamate or
lysine) can catalyze the thermal copolymerization of amino acids. You don't
even need high heat. Thermal protocells have been made at temperatures as
low as 70 C in a few days in the presence of phosphoric acid or in a few
months in the absence of phosphoric acid. The only requirements for making
thermal proteins is a source of heat, anhydrous conditions and the presence
of a dicarboxylic amino acid, lysine or possibly glycine in at least
equimolar amounts to the other amino acids. The only requirement for making
protocells is cooling and rehydrating the thermal proteins.
>
> Clearly, there are several relevant follow-up points to the
> claims that you make.
>
I would point out that for this model all claims are based on experimental
evidence rather than theoretical speculation.
>
> 1. Where did the nucleotides come from?
>
For experimental purposes nucleosides were provided for the protocells to
absorb and utilize. The protocells then converted them into nucleotides
using light and either ATP or pyrophosphate, whatever was provided
(protocells provided with nucleosides and pyrophosphate tended to make ATP as
well). Abiogenically, while it is more difficult to make bases, sugars,
nucleosides or nucleotides than it is to make amino acids, it can be done
well enough to provide protocells with the necessary raw materials. However,
this is research that is yet to be done.
>
> 2. As for these nonrandom sequences that are catalytically active,
> what sequences are we talking about?
>
I'm not sure I understand your question. If you are asking me what the
specific sequences are, I cannot tell you, since to my knowledge no thermal
protein has been sequenced yet. This is not too unusual; I've worked on some
cellular proteins that proved to be so difficult to sequence they still
haven't been even some years later. Nonrandomness has been established by
other means, namely that thermal proteins produce only a few descrete bands
or peaks during purification procedures, the amino acid composition of the
thermal proteins differs from that of the starting mixtures, the same
mixtures produce the same thermal proteins based on amino acid composition,
catalytic activity and the sequencing of hydrolysis peptides (the same
mixtures produce thermal proteins with the same amino acid compositions, the
same catalytic activity and the same peptide sequences), and the catalytic
activity is significantly greater than that of the original mixture of amino
acids, can be inhibited and recovered, and displays Michaelis-Menton kinetics.
>
> What biological proteins have these sequences?
>
First of all, let me make sure you are not laboring undering a few
misconceptions. Thermal proteins do not need to be identical to contemporary
cellular proteins to be the precursors of contemporary cellular proteins.
Nor is it required that thermal proteins evolve into contemporary cellular
proteins. All that is necessary is that thermal proteins perform the same
functions as contemporary cellular proteins and help to produce a system that
can eventually evolve to produce contemporary cellular proteins.
Having said that, there is evidence that some sequences found in contemporary
cellular proteins are fairly ancient, and the thermal protocell model
predicts that the direct ancestors of contemporary cellular proteins were
polypeptides produced by protocells using as templates polynucleotides that
they created using thermal proteins as templates. So it is not inconceivable
that a few thermal protein sequences could have found their way into
contemporary cellular proteins. I would not expect it, however, and until
thermal proteins can be regularly sequenced we won't be able to see if any
thermal protein sequences are in contemporary cellular proteins.
For now, a better test would be to see if thermal proteins could be created
that mimick the activity of contemporary cellular proteins. At least one
such test has been done successfully. When the amino acids that form the
active site of melanocyte-stimulating hormone (MSH) -- glutamate, glycine,
histidine, arginine, phenylalanine, and tryptophan -- are used to create
thermal proteins, the resulting polymers displayed the activity of the
hormone, though it was weaker. However, polymers that were made from
mixtures that lacked even one of the critical amino acids (such as arginine)
displayed no hormonal activity whatsoever. This activity can be inhibited by
the same agents that inhibited the hormone (agents like norepinephrine), and
the same agents that restore function to the hormone also restore function to
the thermal proteins. As such, it would appear that thermal proteins can
reproduce the activities of contemporary cellular proteins. How they do so
is still uncertain, but thermal proteins tend to be large enough (> 5000 Da)
that some form of three-dimensional structure is probably involved.
>
> 3. What percent of the biological protein modules are explained
> as being derived from thermal proteins (after all, most protein
> modules are very old)? Has anyone resolved the 3-D structure
> of a thermal protein?
>
Very few such attempts have been done; in once case with one amino acid
mixture no evidence of helicity could be found. Considering that L amino
acids tend to racemize when heated, and that known secondary structures
require homostereoisomers, it is unlikely that thermal proteins reproduce the
same kinds of structural/functional domains found in contemporary cellular
proteins.
Again, however, they are not required to, either to be functional or to be
precursors of contemporary cellular proteins. Secondary structures and
structural domains are known simply to stabilize the shape of cellular
proteins. In fact, they are required, because cellular proteins contain only
a few amino acids that are used in the active or regulatory sites and often
they are spread out along the primary sequence, so the cellular protein has
to fold just the righ way to bring all the active/regulatory site amino acids
together in just the right orientation in just the right places to be
functional. The secondary structures and the domains then ensure that this
folding remains stable, while at the same time allowing enough give and take
to permit shifts in conformation (overall shape) in response to substrate or
chemical regulator binding. These active/regulatory site amino acids are in
fact mainly found in what are generally called the random coil sections of
the protein; in other words, they are generally not part of a secondary
structure, though they are often part of a domain.
In contrast, what we know of thermal proteins so far suggests that the active
site amino acids are more numerous and closer together, hence a precise shape
may not be necessary. I suspect that thermal proteins are composed of short
repeating sequences that allow the active amino acids to cluster together.
As a result, a thermal protein may in fact have several "active sites". Some
local folding may be necessary to group the clustered amino acids in the
correct orientation (forming protodomains), but the rest of the thermal
protein is probably one huge random coil. But until we can isolate a single
polypeptide (or create one synthetically using a sequencer) methods such as
x-ray crystallography and circular dichroism (which can give accurate
information of shape) will be largely useless.
>
> 4. The process of converting light into ATP is known as photosynthesis
> in living cells.
>
Of course.
>
> Only two main processes exist - the photosystems of
> eubacteria (which employ an elaborate network of precisely positioned
> chlorophyll molecules and Fe-S clusters) and bacteriorhodopsin, which is
> found in one species of archaebacteria (and functions like vertebrate
> rhodopsin).
>
Correction: only two main processes **are currently used** by contemporary
organisms. That does not mean there were never any other processes that got
weeded out by evolution as too inefficient or were lost when the organisms
that used them became extinct for other reasons.
>
> The thermal protein system is *homologous* to which one?
>
Again, this question presupposes that contemporary photosynthetic processes
are derived directly from protocellular processes, when in fact conteporary
processes almost certainly evolved from simpler **cellular** (rather than
protocellular) processes. Remember, protocells were most likely analogous
structures that eventually evolved into simple homologous structures;
contemporary photosynthetic processes probably directly derive from this
point rather than some previois point.
I am not yet aware of any proposed mechanism for how protocells perform
photosynthesis, but there are two possibilities. The first is that
structures known as flavins and pterins are created during the thermal
copolymerization process and these associate directly with the thermal
proteins. The second is that porphyrins are rather easy to make abiotically
and are quite stable. They have been demonstrated to be readily incorporated
into thermal protocells, and of course chlorophyll is at heart a porphyrin.
Regardless of what served as the light-capturing agent, the most plausible
mechanism would probably go like this: an exciteable ring-based structure
absorbs light; this in turn excites an electron which jumps out of the
structure and is captured by a lysine-rich thermal protein; this protein uses
these electrons to bind pyrophosphates to nucleosides picked up from the
environment; some of these nucleosides are converted into
triphosphonucleotides like ATP while others are simply converted into
monophosphonucleotides. Other thermal proteins can use these electrons to
perform catalytic activities, such as decarboxylation.
>
> These questions seem very important, as I think most origin-of-life
> researchers view thermal proteins as interesting chemistry in-of-itself,
> but not very helpful in explaining the origin of the life forms that
> exclusively dominate this planet....
>
This has more to do with their professional biases for alternative theories
than with any empirical or theoretical problems with thermal protocells.
>
> ...(i.e., proteinoids might be somewhat
> analogous, but are they homologous to life forms?)
>
Again, this persupposes that the first truly living cellular structures must
have been simpler versions of contemporary cells. One thing that virtually
all abiogenecists agree upon is that in fact the first living cellular
structures used (to use your tems) analogous structures and processes, then
evolved into the homologous common ancestor of contemporary life. Most
people who reject thermal protocells do so, not over concerns of analogous
vs. homologous, but because they believe genes came before proteins, or
protocells had to have lipid bilayer membranes, or they object to the
nonrandom nature of the thermal proteins, or they believe the environment of
the early earth was incompatable for thermal protein formation, or etc.
>
> >The information for doing all this comes from the amino acids and the
> >resulting thermal proteins themselves, from their physiochemical
structure
> >and function. Davies couldn't bring himself to accept that the answer
> >could be so simple, yet in science often the most profound mysteries turn
> >out to have very simple answers.
>
> Clearly, protein function follows from structure, which in turn follows
> from the primary sequence of amino acids. But if this answer and
> process is this "simple," why is life monophyletic?
>
Because contemporary life had only one common ancestor.
>
> Why haven't new
> life forms emerged many times over in the past 4 billion years?
> Shouldn't we be able to find very different life forms existing today
> that arose 1.5 billion years ago, 0.85 billion years ago, etc.? We don't.
> Why not?
>
One reason is that an oxygen-rich atmosphere prevents the kind of abiotic
reactions that create large amounts of amino acids; with no abiotic source of
amino acids there can be no thermal copolymerization of polypeptides and thus
no protocells. Another reason is that life began to find ways of making its
own supply of micromolecular precusors like amino acids. Once this
supplanted abiotic sources, it was no longer possible to create
concentrations of amino acids in a free state; instead, amino acids became
almost exclusively tied up as cellular protein in living organisms. Another
reason is that contemporary lifeforms -- especially bacteria -- would tend to
consume any abiotically generated micro- or macromolecular precursors before
they had a chance to form cellular structures. Another reason is that
protocells themselves make excellent bacteria food, especially if they were
made out of protein.
The point is that once the environment of the earth changed to the point that
abiotic processes could no longer occur and once contemporary life became
dominant, no new form of life would have the chance to get started much less
become established.
>
> >The secret is that the complexity and organization of the universe
> >is built up, layer by layer, from simple beginnings that have the
> >ability to arrange and rearrange themselves in tremendously varied
> >combinations. In other words, the information for life is already
> >built into the universe when it first appears.
>
> As I mentioned, I do indeed see the theological appeal behind this
> suggestion and I cannot say it is wrong.
>
And as I explained earlier, I find no theological appeal in this model. The
appeal for me is that it is the only model to have such strong experimental
support, and it is the only model to have created anything like a living
cellular organism in the lab; for me the appeal is entirely empirical.
>
> But you seem to be overlooking some things.
>
Most of the following objections are based on certain misconceptions on your
part, not the least of which is the assumption that protocells must have been
homologous with contemporary cells in order to be evolutionary precursors.
>
> Biological proteins hook up primarily through classical
> peptide bonds, yet I seem to recall that the amino acids in thermal
> proteins hook up in various ways. Thus, what would be the informational
> source that specifies the use of only peptide bonds, especially when
> thermals can do all the things you mention?
>
In point of fact there is only one piece of evidence that suggests that
thermal proteins contain other kinds of bonds besides peptide bonds. A
researcher used the dye biuret, which binds almost exclusively to peptide
bonds, and discovered that thermal proteins bound only about 50% the amount
of dye per dalton as cellular proteins. Unfortunately, biuret is itself a
rather insensitive assay, and tends to give different results even among
cellular proteins. Biuret also cannot bind to non-alpha-peptide bonds, so a
thermal protein made up of 100% peptide bonds of which half are alpha and
half are epsilon would produce the same results. Besides, NMR and IR
spectroscopy -- which can identify the different bonds -- have shown
conclusively that thermal proteins are virtually entirely composed of peptide
bonds.
They can have mixtures of different types of peptide bonds, but that depends
upon what amino acid is being used to catalyze the polymerization. Thermal
proteins based on aspartate tend to have roughly 50% alpha bonds and 50% beta
bonds, whereas those based on glutamate have virtually 100% alpha bonds.
Lysine-rich thermal proteins have some epsilon bonds, but they tend to be a
third or less.
Again, however, the question is not how did thermal proteins evolve into
cellular proteins, but how could thermal proteins help to give rise to
cellular proteins? The simplest explanation based on the experimental
evidence seems to be that the thermal proteins served as templates for making
polynucleotides, which in turn served as templates for making polypeptides.
Before the advent of tRNA, the amino acids and the polynucleotides bound
together directly. (There is evidence to show that amino acids bind
preferetially to the same codon dublets that make up the basis for the
genetic code: phenylalanine to UU; lysine to AA, proline to CC and glycine
to GG for example.) Research shows that the most stable arrangement of amino
acids permits only alpha bonds to occur. Also, protocells tend to create
polyaminoacyl adenylates (which can be used as templates for making either
polynucleotides or polypeptides), which also permit only alpha bonds to be
used.
In other words, the switch to exclusive use of alpha-peptide bonds probably
occurred when protocells started making polypeptides from polynucleotide
templates, and the source of the information for this switch was the
physiochemical interaction of amino acids to nucleotide dublets.
>
> Secondly, biological proteins
> use only a specific set of 20 amino acids. Do thermal proteins
> only form from this set or can they form from the hundreds of other
> types of non-biological amino acids also? If they can only form from the
> set of 20, your case is more appealing. If they can form from any
> amino acid, you are missing another layer of specification.
>
Thermal proteins can form from nonproteinous amino acids, but that doesn't
complicate the scenario. First of all, there is experimental evidence that
demonstrates that nonproteinous amino acids do not bind to nucleotides
dublets as well as proteinous amino acids, as such it is unlikely that the
polypeptides formed from polynucleotides would incorporate anything other
than proteinous amino acids. Secondly, many of the nonproteinous amino acids
have R groups that create too much steric hindrance to permit proper folding.
While this would not be a serious problem for thermal proteins, it would be
a potentially fatal problem for cellular proteins, so any nonproteinous amino
acids that had started out as proteinous but which prevented the formation of
properly functional proteins, could have been weeded out by natural
selection. The redundancy of the genetic code suggests that something like
this happened more than once.
>
> I find your scenario to be interesting, but I think you are
> overstating your case.
>
On the contrary, based on the amount of evidence I've read, I believe that if
anything I am understating my case. There are still problems and
uncertainties that need to be addressed, but I believe their resolution will
only strengthen the case for thermal protocells.
>
> The origin of life is not a problem of
> coming up with "information", it's about explaining the origin
> of the information that exists
>
And that origin is explained by the physiochemical interaction of micro- and
macromolecular precursors, which serves as the basis for self-organization.
Kevin L. O'Brien