Antibody Enzymes and Sequence Space


(Ann Gauger) #41

@swamidass I don’t know if you saw this down below but I think it’s brilliant, If you are confident of this 10^-10 frequency for obtaining beta lactamase activity, even better than wild type, then here’s a way to prove it.

@Rumraket, you gave me the idea. If you are sure of the abzyme result then go into the lab, prepare large trays, lots of them, with ampicillin. It has to be fresh because it loses half its activity every day. Then plate about 5 x 10^10 bacteria that lack resistance to ampicillin. You can start with a half dose if you want. What do think? Will 5 of them spontaneously discover a new way to break down ampicillin? No?

Gosh, there have to be a bunch of enzymes from the beta lactamase structural family in there already. Surely one of them…

Well, it will be tricky proving it’s not a contaminant, so use really good sterile technique, and be ready to determine sequence and structure for a beta lactamase!

Oh wait. If it was this easy. somebody would have done it already. Does that tell you anything?

(S. Joshua Swamidass) #42

Um, I thought that precisely this has been done already. What am I missing?

(Ann Gauger) #43

It should be reproducible in cells.

(S. Joshua Swamidass) #44

And it is…? I’m really lost on your point. It seems like you think that experiment hasn’t been done.

(Ann Gauger) #45

No, my experiment hasn’t. Not in cells, not the way it would have to happen in evolution.

(S. Joshua Swamidass) #46

Okay, if you say so. I suppose I can show you otherwise sometime this weekend if we have time. :man_shrugging:

(Ann Gauger) #47

You can’t show me something that isn’t there. Maybe I’ll show you.

(S. Joshua Swamidass) #48

Also, to be clear, the experiment does not show the abzymes are better than wildtype. That is misreading the results.

(Ann Gauger) #49

I agree. I thought I heard you say that though.

(S. Joshua Swamidass) #50

Nope. Not at all. I said they were better than wildtype at a specific concentration, but we don’t know the concentration of the abzymes, so it isn’t on the same scale.

Your interpretation of that paper is hard for me to understand. I understand you don’t want to concede any points on Axe’s argument publicly. Fine. I wonder if stepping through that paper in person, off the record, might bring some more clarity to your reasoning. Speaking bluntly, I really can’t follow your logic here, and usually I can. It makes me wonder if there is an important piece of the puzzle one of us is seeing and the other is not. Maybe it will be easier to make sense of it off the record.

(Ann Gauger) #51

Personally, I don’t know why you can’t see that there is a big disconnect with these reported results and our experience in the lab and in medicine. If it was so easy to get beta lactamase activity penicillin would have been a dismal failure as an antibiotic decades earlier. The numbers don’t make sense. I can see why you would want Doug to be proven wrong. But to ignore the ridiculousness of what a frequency of 10-10 would mean is not like you.

There are about 100 trillion bacteria in our microbiome one estimate says.
10^14 cells. That means there should have been nascent ampicillin resistant cells on or in each and every one of us, from the beginning of the use of the antibiotic. Yet penicillin was an effective antibiotic for quite a long time.


(S. Joshua Swamidass) #52

Hmm…there are and were nascent amp resistant cells in most people, even before exposure to amp. Perhaps there are some misunderstandings here about the nature of clinical antibiotic resistance…

Come to think of it, the same misunderstanding seems to be in Behe’s malaria logic…

(Mikkel R.) #53

5x10^10 different bacteria =/= 5x10^10 different protein sequences.

If I plate a colony of bacteria, I’m going to plate mostly clones, differing by about a single substitution in roughly every 10 indviduals, give or take. And since breaking down ampicillin is not the only way for a bacterium to deal with the challenge of it’s presence, even under multiple rounds of plating and selection I will not be guaranteed to be selecting for enzyme activity, as I could equally well be selecting for a more discriminating transporter, or a host of other possible solutions.

I’m sorry but the experiment you propose would not actually constitute a test of the hypothesis. We’d need some way to generate several billion different protein sequences. Such experiments have been done, usually using highly mutagenic PCR to generate the different mutants, and then some selection protocol, and they frequently find the functions of interest.

But if the conclusions sold to the averge IDcreationst based on Axe’s work was true, such results should be impossible to obtain, because the average ID creationist out there believes Axe has shown that functional proteins exist at an average rate of 1 in every 10^77 protein sequences. And last I checked, we haven’t been able to screen a sphere of proteins the size of the solar system, yet functions are still reliably found.

(Mikkel R.) #54

10^14 cells =/= 10^14 different proteins.

You yourself seems to understand this distinction here:

There’s a difference between saying “an enzymatic function exists at a rate of approximately 1 in 10^10 proteins” and “that level of diversity of different proteins exist at any given moment in some bacterial population of a few trillion, most of which will be clones”. To test the claim that the function is found at a rate of ~1 in 10^10, we’re going to need to actually generate those 10^10 different proteins and test if the function is present. The key word in different proteins is different. Functional proteins are generally found clustered together(if a given protein has some function, chances are nearby proteins also has some level of that function), so that would imply for every one such functional cluster, there’s about 10^10 more space without that function. That means we have to sample more broadly than just doing 10^10 variants of almost identical proteins to get further out and sample into other potential clusters if they’re out there.

That takes time we don’t have if we just let a natural population incrementally generate that diversity, so we have to speed up the process of generating diversity. Diversity that a natural population could generate over a geological blink of an eye, but which is still well beyond what is practically feasible for human scientists.

(Bill Cole) #55

Impossible to obtain is a meaningless statement. The question is about the mechanism of RMNS producing the result. In Axes case he shows how many trials are required to build a substrate and an active cite in a living organism.

(John Mercer) #56

I just want to ask, again, why you are pretending that this is the only catalytic antibody paper you need to deal with. There are more than 5000 that you are ignoring. Why?

(John Mercer) #57

How many of the >5000 catalytic antibody papers have you read?

Why do you persist with this false portrayal of one paper against one paper? Scientists do not read the literature in that way.

(Mikkel R.) #58

Heh, no it’s actually very concrete. But I will concede it’s an overstatement. Impossible implies it can’t happen, full stop. While Axe’s numbers of course merely implies it would be incredibly unlikely.

The question is about the mechanism of RMNS producing the result. In Axes case he shows how many trials are required to build a substrate and an active cite in a living organism.

No, he doesn’t. At all. I have a hard time seeing how even Axe would agree with that. It isn’t clear what you mean by a “trial”. And why would the experiment need to “build a substrate”? The substrate is the compound on on which the enzyme works. The enzyme doesn’t “build” the substrate, it acts on it. It catalyzes the chemical change of it. And his work also doesn’t show how difficult it is to build “an active site”. It is purported to show how rare particular folds carrying out particular functions are. You could merely have read Ann Gauger’s post up above:

(John Mercer) #59

Your failure, and at this point refusal, to read the literature is incredible.

Catalytic antibodies come from the immune system, a completely natural system for looking for something that works from random sequence (V regions).

Here’s one of the two original papers from 1986. It’s a monoclonal antibody that is soluble. None of your objections apply. It took me less than 5 minutes to find a PDF:

Here’s what I don’t get: if someone points out that you’ve ignored a 32-year-old, entire field, wouldn’t you seek out the initial paper(s) on your own?

(John Mercer) #60

Even worse for @Agauger, it is a far better test than Axe’s without even having that as its rationale. And then there are the >5000 other papers she is ignoring.