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Author
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Topic: Back to the Topo II - where does it end?
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Nel
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Member # 614
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posted 12. June 2003 23:17
Nic,
I didn't "express doubts" about Blocker's model. I simply showed that you called an imaginary model a "confirmed prediction". An imaginary model can in no way, shape, or form, be a "confirmed prediction".
Anyway, I'll read it as well as do an RBH and say: What Mike Said! ![[Smile]](smile.gif)
If I have any more comments on that paper I'll return.
[ 12. June 2003, 23:45: Message edited by: Nelson_Alonso ]
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yersinia
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Member # 324
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posted 13. June 2003 01:36
Well, how fast the tides turn. Earlier in this thread, I had quoted Blocker et al.:
quote:
The similarity of the beta subunit to FliI is something like 33% which is highly statistically significant, well above the ambiguous level:
quote:
How might these ATPases catalyze processive protein export? Spa47 (the Shigella FliI homolog) shares 33% amino acid identity with the beta-subunit of F1-ATPase. Proteins with >30% sequence identity have a high probability of sharing similar structures (69). Active F1-ATPase is a heterohexamer consisting of alternating alpha- and beta-subunits with a gamma-subunit inserted in a central channel where it rotates during the catalytic cycle (70). No equivalent of the alpha-subunit of F1-ATPases is found within flagellar or TTSS-encoding operons, so we assume that the type III export motor is a homohexamer. When modeled on the F1 structure, Spa47 fits at the inner membrane base of our NC structure (Fig. 3). It would contain a central channel aligned with the one found within the NC and of similar diameter to it, through which the proteins could be secreted (see Supporting Text).
Source: Blocker A, Komoriya K, Aizawa S. Proc Natl Acad Sci U S A 2003 Mar 18;100(6):3027-30. Type III secretion systems and bacterial flagella: Insights into their function from structural similarities. http://www.pnas.org/cgi/content/full/100/6/3027
...the assumption of homology seems to be confirmed by the fact that the resulting protein complex fits well into their model of T3SS structure.
In this quote they:
- 1. Note the statistically significant sequence similarity between FliI/Spa47 and beta-F1-ATPase
- 2. Based on this evidence of homology, make a prediction about the structure of Spa47, and
- 3. Test it by placing the inferred structure into the base of the T3SS (width and configuration derived from electron microscopy etc.) and
- 4. Note that it just happened to fit, even though it was perfectly possible that the structures would have been the wrong size to fit. This strengthened the inference of homology.
This is what Nelson apparently disputed:
quote:
Although the point of FliI homology seems moot upon careful reading of the paper you cite:
Source: Blocker A, Komoriya K, Aizawa S. Proc Natl Acad Sci U S A 2003 Mar 18;100(6):3027-30. Type III secretion systems and bacterial flagella: Insights into their function from structural similarities.
I read the paper last night, and it seems you are quite wrong about this. There is no evidence for a Spa47 protein complex, a fliI that is homologous to the b subunit of the F-ATP synthase. What is shown in green in fig 3 in the paper is only an imaginary model of Spa47 based on the F1 structure and that model can be successfully docked to the real NC outline where they would imagine it would have to sit to pump secrete proteins into the channel. The evidence here is pretty weak.
[...]
...much like your Spa47 blunder...
[...]
Everytime you show me a list of citations, the first few turn out to be a wild goose chase, as with the Spa47 paper.
[...]
Nic, there is no such thing as a protein complex of Spa47. It's ficticious, imaginary, like Santa Clause, the Easter Bunny, Pumpkin Head, the Matrix, X-men, spider-man, super-man, echo and the bunnymen(actually they exist). It's made up. Of course it fits, the whole thing is imaginary. How can it be a confirmed prediction? Once the post-doc finds it (or something like it) then the prediction that the model makes can be confirmed.
The question is important, because if homology is conceded, then Nelson can no longer claim that there is no evidence for cooptable structures preceding the IC structures like the flagellum (e.g. in this thread), and he can no longer defend Dembski's calculation as actually taking into account the evidence for precursors:
quote:
As I already showed, Dembski's calculation is not like assembly a plane through a tornado in a junkyard. It is precisely attempting to find a forward-chaining of events that would have selection as the driving force all the way up mount improbable.
.
I raised (here) other examples of important facts that Dembski calculation didn't take into account, for example ExbBD-MotAB homologs, internal homologies of the axial proteins (rod-hook-linkers-filament), and analogies showing how various intermediate non-flagellum stages could still be functional.
Much of the thread was spent debating these various points (people can judge for themselves). In my opinion it is clear that Nelson's ID reasoning led him astray repeatedly:
- 1) For awhile he argued strenuously ("near certainty") for the external-proton-channel model of MotAB's motor mechanism. Evolutionary considerations and homology with ExbBD indicate an internal channel, and in fact all of the most recent papers appear to now endorse the internal channel model.
- 2) Nelson argued that "simple" pili had to have 5 parts in their extracellular filament, but the Hrp pilus apparently has only 1-2
- 3) Nelson argued that nonflagellar filaments had to be assembled from the base rather than the tip, but this was also disproven by the Hrp pilus and other Type III pili.
- 4) Nelson argued that the distribution of ExbBD was more restricted than the distribution of MotAB, indicating that ExbBD was derived, but the existence of ExbBD but not MotAB in cyanobacteria and archaea disproved that.
- 5) Was his assertion that the evidence was weak for the homohexamer model for FliI/Spa47. And yet, the Blocker et al. proposal, clearly based on common evolutionary reasoning, appears to have won out here also.
But now we have Nelson saying that he wasn't expressing doubts, and Mike Gene saying he wished he'd predicted the homohexamer model, and Nelson saying "what Mike Gene said"!
(even though MG's flagellum essays were clearly the source of Nelson's claims #1 and #2, perhaps #3, and IIRC MG tried out #4 on ARN a few months before Nelson tried it here!)
I'm not sure we could have a clearer case of ID thinking consistently leading to the wrong answers on a variety of specific empirical questions.
[edits for format etc.]
[ 13. June 2003, 01:44: Message edited by: yersinia ]
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yersinia
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posted 13. June 2003 02:09
MG writes:
quote:
The truly interesting stuff comes from the FliI and F ATPase subunit comparison. The essence of their similarity revolves around the ATP-binding domain. At first glance, this doesn’t seem too useful in distinguishing between common design and common descent, as one might expect ATP-binding motifs to be reused in different contexts from either perspective. But what seemed to indicate a hodgepodge use of this motif was that it appeared to be used as a monomer in the flagella. There didn’t seem to be any type of design logic behind its reuse.
Your logic is none too clear here, but you seem to be under the misapprehension that the similarity between F1-beta and FliI was thought to be just a matter of the ATP-binding motif. This is not what I've gathered:
quote:
The proteins needed for flagella rotation include one that has antibody reactivity like and extensive sequence homology with the ATP synthase beta-subunit (243)
(Boyer 1997, Ann Rev Biochem, "The ATP synthase -- a splendid molecular machine, p. 743)
Ref #243 is:
quote:
J Bacteriol. 1993 May;175(10):3131-8.
Genetic and biochemical analysis of Salmonella typhimurium FliI, a flagellar protein related to the catalytic subunit of the F0F1 ATPase and to virulence proteins of mammalian and plant pathogens.
Dreyfus G, Williams AW, Kawagishi I, Macnab RM.
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06511-8148.
FliI is a Salmonella typhimurium protein that is needed for flagellar assembly and may be involved in a specialized protein export pathway that proceeds without signal peptide cleavage. FliI shows extensive sequence similarity to the catalytic beta subunit of the F0F1 ATPase (A. P. Volger, M. Homma, V. M. Irikura, and R. M. Macnab, J. Bacteriol. 173:3564-3572, 1991). It is even more similar to the Spa47 protein of Shigella flexneri (M. M. Venkatesan, J. M. Buysse, and E. V. Oaks, J. Bacteriol. 174:1990-2001, 1992) and the HrpB6 protein of Xanthomonas campestris (S. Fenselau, I. Balbo, and U. Bonas, Mol. Plant-Microbe Interact. 5:390-396, 1992), which are believed to play a role in the export of virulence proteins. Site-directed mutagenesis of residues in FliI that correspond to catalytically important residues in the F1 beta subunit resulted in loss of flagellation, supporting the hypothesis that FliI is an ATPase. FliI was overproduced and purified almost to homogeneity. It demonstrated ATP binding but not hydrolysis. An antibody raised against FliI permitted detection of the protein in wild-type cells and an estimate of about 1,500 subunits per cell. An antibody directed against the F1 beta subunit of Escherichia coli cross-reacted with FliI, confirming that the proteins are structurally related. The relationship between three proteins involved in flagellar assembly (FliI, FlhA, and FliP) and homologs in a variety of virulence systems is discussed.
Finally, in support of the contention that the similarity is in excess of what can be accounted for simply by both proteins being ATP-binders, looking at Vogler et al.:
quote:
Variants of motifs A and B are present in many ATP-utilizing enzymes. However, the degree of agreement between these motifs in FliI and the F0F1 and related ATPases is generally stronger than that between FliI and the ATP-utilizing proteins generally...We conclude that FliI is specifically related to subunits of the F0F1, vacuolar, and archaeabacterial ATPases and not just generally related to nucleotide-binding or nucleotide-utilizing proteins as a whole.
(p. 3569, Vogler et al 1991, Jour. Bacteriology)
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Mike Gene
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posted 13. June 2003 02:41
Yersinia: Finally, in support of the contention that the similarity is in excess of what can be accounted for simply by both proteins being ATP-binders, looking at Vogler et al.
Sure. But as your recent find illustrates, its not a question of simply being an ATP-binder. The design logic goes further.
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Nel
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Member # 614
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posted 13. June 2003 23:36
Unfortunately, I have to quote Nic for context, but I have tried to keep the quoting to a minimum.
First of all, I have been extremely consistent about what I think of Blocker's model and what I think about your misinterpretation of it. How can their assumption be confirmed if the resulting protein complex is completely imaginary? It can't be confirmed until they find the protein complex.
In your post you make this claim.
quote:
4. Note that it just happened to fit, even though it was perfectly possible that the structures would have been the wrong size to fit. This strengthened the inference of homology.
I noted all of this already in the post in this thread dated May 2003 15:16. For some odd reason you think I disputed this.
quote:
What is shown in green in fig 3 in the paper is only an imaginary model of Spa47 based on the F1 structure and that model can be successfully docked to the real NC outline where they would imagine it would have to sit to pump secrete proteins into the channel. The evidence here is pretty weak.
The evidence is pretty weak, they don't really know if their imaginary model is real, or if it really can succesfully dock to the real NC outline, it probably can, but there isn't a confirmed prediction. It would be a confirmed prediction if they actually find the protein complex that is based on the F1 structure of the F-ATP synthase and it actually sits on the needle complex secreting proteins. It would be even better for them if they find a protein complex that closely matches theirs. The homology inference cannot be confirmed until the protein complex is found. But then, what would be the point? Why do I say this? Let me explain. This is closely related to Mike's post, which is why I highly endorsed it.
You add that the prediction that Mike makes somehow is an indication of a tide turning. And yet I made exactly the point that Mike made almost a month ago in my summary post:
quote:
However, each of these three systems are very different from eachother, not only that, the type III secretory system most likely evolved from bacterial flagella
So the whole thing is moot anyway. It is simply not true that if homology is conceded, then I "can no longer claim there is no evidence for cooptable structures preceding ... the flagellum". Why not? Well, for starters, type III systems postdate the flagellum! There are a great many reasons to think this, which is why even the literature is converging on this line of thought:
Phylogenetic analyses of the constituents of Type III protein secretion systems, J Mol Microbiol Biotechnol 2000 Apr;2(2):125-44
And that is what the Blocker paper is discussing, type III systems.
Yersinia writes:
quote:
I raised (here) other examples of important facts that Dembski calculation didn't take into account, for example ExbBD-MotAB homologs,
Let me remind you that there is no such thing as an ExbBD-MotAB homolog, there is very weak similarity only between ExbB and MotA, thats it.
quote:
internal homologies of the axial proteins (rod-hook-linkers-filament),
There is a paper that discusses this:
quote:
Some degree of similarity among the sequences of the axial proteins would not be surprising, for two reasons. The first is that, since the axial proteins together form a continuous filamentous structure, we might expect them to have a similar lattice and to share common structural elements that determine the lattice....The second reason for looking for sequence similarities among the axial components concerns the manner in which they are thought to be exported across the cell membrane.
Homma, M, DeRosier, DJ, and Macnab, RM. 1990. Flagellar hook and hook-associated proteins of Salmonella typhimurium and their relationship to other axial components of the flagellum. J. Mol. Biol. 213: 819-832.
In other words, the similarities here can be explained in functional terms, and does not necessarily mean that they all evolved from simpler precursors.
quote:
and analogies showing how various intermediate non-flagellum stages could still be functional.
Ok show me how various intermediate non-flagellum stages could still be functional. Lets start with alternative functions of the export machine, or from the m ring to the rod proteins.
Let me now respond to each of Nic's numbered points. You can refer to them above, I don't want to quote because it's cluttering up my post.
First, there is no need to argue strenuously for an external-proton-channel model of MotAB, not even Blair's model contradicts an external-proton-channel model for the motor of flagella.
Furthermore, not even Blair mentions an "internal" channel for ExbB. So I'm not sure where you get the idea that all papers now endorse an internal channel. In fact, tonB may be involved as either part of the channel or in opening the channel in Blair's model. Nothing in Blair's model, nor in the ExbBD complex itself, requires that the channel is internal.
With respect to the required components of pili, what I said was that P type pili require 5 parts to function, as well as the needle complex of the T3SS. What Nic is calling the "hrp pilus" is the external needle. However, even so, it's still not composed of just 1 or 2 parts. In fact, research is constantly uncovering more parts to the the hrp system:
quote:
Using a polyclonal antibody against partially purified Hrp pili, we have detected three additional proteins associated with Hrp pili: two of approximately 68 kDa in size and one of 49 kDa. These three proteins were detected in the Hrp pilus preparation from Pst DC3000, but not from the hrcC, hrpS or hrpA mutant. We are investigating whether the three proteins are additional subunits of the Hrp pilus.
http://www.bspp.org.uk/icpp98/1.7/5S.html
What we actually know from that forms the pilus is HrpA, HrpZ, HrpW, and AvrPto, 5 parts. Hrp pilus doesn't have only 1-2 parts, although one part, HrpA, seems to be the main structural component, there is obviously a lot more to the story than that. In fact, I can't find a 2-part pilus anywhere. Not even in type III secretory systems. Even if I did find it on T3SS, what would be the point? We are almost sure where that came from anyway.
quote:
3) Nelson argued that nonflagellar filaments had to be assembled from the base rather than the tip, but this was also disproven by the Hrp pilus and other Type III pili.
What I actually said was that P type pili were not built from the bottom up, like flagella. So this was not disproven by type III secretory systems, since I was not discussing type III secretory systems, which there are good reasons to think that they evolved from flagella. So that type III secretory systems are built from the bottom up is no surprise. What would be surprising is if P type pili were built that way, they aren't.
With respect to ExbB, what you showed me was non-flagellar homologs of ExbB and MotA in a tree that was based on a hypothetical protein. The problem with your excercise is that true ExbB proteins are all based on the function of ExbBD, which is restricted to the gram-negatives.
I have actually done quite a few excercises here which lead me to find out what Nic's error was, for anyone wanting to do this on their own, one way to phrase the question is, how many regions on the tree have authentic Mot proteins but do not have authentic Exb proteins, and how many regions of the tree have authentic Exb proteins but do not have authentic Mot proteins?
In order find real Mot proteins, you would need to find other flagellar proteins in the same species, and conservation of certain key residues. Same for Exb.
In fact, I doubt that you can show me an authentic ExbB protein in any gram-positive bacteria. But I can find MotA in both gram-positive and gram-negative bacteria. In other words, I can always find more MotA proteins then you can find ExbB proteins, because ExbB proteins are restricted to gram-negatives. It's as simple as that.
[ 13. June 2003, 23:47: Message edited by: Nelson_Alonso ]
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