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The Talk.Origins Archive: Exploring the Creation/Evolution Controversy

Nylonase Enzymes

Post of the Month: April 2004


Subject:    Re: Nylon bug
Date:       20 April 2004

G'Day All

On Tue, 13 Apr 2004 19:33:51 +0000 (UTC), Black wrote:

>AiG has a new article about the nylon bug on their site:
>I'm not a biologist, so I don't know anything about this. What do you think?

As usual, it is typical AiG misunderstanding and misinformation.

First, a little background. Nylon is an artificial polymer not found in nature. Indeed, not only is the nylon polymer not found in nature, neither are the linkages that bind the subunits together. Nylon first entered the environment in the 1930's. By 1975, bacteria capable of hydrolysing nylon were found in wastewater from nylon plants.

Now, it is obvious that the gene(s) for hydrolysing nylon cannot have been present from the beginning, as in the absence of the substrate (nylon, not present before 1930), the gene product is non-functional, and the gene would be mutated to uselessness (or an entirely different function) in a few hundred years by random mutations, let alone thousands.

Faced with such an obvious production of a new gene with a novel function, the first thing creationists tried to do was claim this was a loss of information, that the nylonases represented a protein-digesting protein (protease) that had lost substrate specificity. That of course didn't fly, as the nylonases are exquisitely specific, act on no known amide bond other than the nylon beta amide bond, and have no relationship to any known protease.

Now AiG is trying to claim that the nylonases are not due to random mutation and natural selection. Their arguments are spurious to say the least.

Some more background, in Flavobacterium there are four nylonase genes, nylA, nylB and nylB' (which are duplicates) and nylC, carried on one plasmid. In Pseudomonas there are two nylonase genes (nyl A and nyl B, homologous to the nylA and nylB genes in Flavobacterium) carried on two different plasmids. The nylB gene does most of the heavy lifting, so to speak, and it is the nylB gene that was formed from a deletion mutation and subsequent frame shift in the RSII repetitive element. The key here is that the mutation was in an internally repetitive sequence of DNA. Frameshifts in non-repetitive sequences usually end up with a high probability of producing a premature stop codon, resulting in production of short non-functional proteins. However, repetitive sequences are very likely to not produce premature stop codons, and it is likely that long, functional proteins can be produced by frameshifts in these proteins. In the case of nylB, an insertion of a T at position 99 in the repetitive sequence resulted in a start codon and a stop codon some 392 amino acids away.

Now onto AiG's claims.

"Evidence against the evolutionary explanation includes: 1. There are five transposable elements on the pOAD2 plasmid."

Transposable elements are relatively short sequences of DNA that can move around the genome by themselves. Don Batten suggests the presence of transposable elements means the plasmid is "designed" to be adaptive. Well, transposable elements can result in rapid adaptation, but the mechanism is pure random mutation and natural selection. Transposons are not specifically targeted anywhere, but jump about at random without regard to the cell's "need". They can generate new enzymes by producing recombination of existing enzymes, but they are just as likely to cause damage. One strain produced by researchers had lost nylA as the transposable elements cut it out. Transposable elements are well known as possible agents of evolution.

"2. All five transposable elements are identical, with 764 base pairs (bp) each. This comprises over eight percent of the plasmid. How could random mutations produce three new catalytic/degradative genes without at least some changes being made to the transposable elements?"

Firstly, the transposable elements are 880 bp and they are not identical, they contain duplications and inversions. Now, Don Batten seems to think that it would take massive mutations to produce the nylonase genes. However, nylB is a one BP insertion, it doesn't take much to make these genes (and nylB' is a duplicate of nylB). Furthermore, the nylA and nylB genes are on different plasmids in Pseudomonas (which doesn't have nylC). It is very likely that the genes arose on different plasmids and were stitched together by the transposable elements at a later stage. Furthermore, the transposable elements (IS6100) are present in many different bacteria and are very strongly conserved, suggesting they do not tolerate mutations very well. So given that the transposable elements are conserved in sequence between different bacteria, and that you don't need many mutations to make a functional nylonase, this objection is void.

"3. All three types of nylon degrading genes appear on plasmids and only on plasmids."

Well, we only HAVE two species of bacteria with these genes, and one seems to have got its genes from another, so it is hardly surprising. They make a big deal that getting all three genes on one plasmid is improbable (but not particularly improbable), while ignoring that in Pseudomonas the (two) genes are on different plasmids and only in Flavobacteria are they are on the same plasmid. Transposable elements have a habit of carrying genes around, so it is not at all unlikely that the genes originally evolved on different plasmids, or even the chromosome, and then were stitched together into one plasmid in Flavobacterium. Furthermore, a large proportion of the genes on plasmids deal with xenobiotic handling or metabolic functions (nylC is next to a cluster of oligopeptide transporters), indeed in Pseudomonas most of the xenobiotic degradation genes are on plasmids, so it is entirely likely that a xenobiotic handling enzyme will arise from mutations of xenobiotic handling genes.

"4. The antisense DNA strand of the four nylon genes investigated in Flavobacterium and Pseudomonas lacks any stop codons."

To start off with, this statement is wrong. NylB, nylB' and nylC in Flavobacterium have no stop codons in the antisense strand, as does nylB in Pseudomonas (it doesn't have nylC). Three of the four genes are nylB, not four independent genes as implied.

Now, having no stop codons in the antisense strand (that is, the partner of the coding DNA strand, remember that DNA is double stranded, and only one strand is translated) is a bit unusual, but the probability they quote (10-12) is dead wrong, the probability is 0.0001. As the three nylB genes are, well, nylB, it is not at all unusual for them to share this property. Furthermore, as nylB is descended from a peptide with many internal repeats, and itself contains a fair number of internal repeats, this makes it less likely to generate stop codons in the first place.

Don Batten's statement, "Yomo et al. also show that it is highly unlikely that any of these genes arose through a frame shift mutation, because such mutations (forward or reverse) would have generated lots of stop codons" is wrong. Yomo et al. show no such thing, they don't mention it at all. As noted above, the precursor sequence to nylB was an internally repetitious sequence, and repetitive sequences are very likely to not produce premature stop codons for significant lengths.

Batten also writes "Some statements by Yomo et al., express their consternation..."

Actually, the statements express their excitement. They think they have found a new evolutionary mechanism (amongst other things they suggest that new genes could be produced from antisense strands of functional genes).

"5. The Japanese researchers demonstrated that nylon degrading ability can be obtained de novo in laboratory cultures of Pseudomonas aeruginosa [strain] POA, which initially had no enzymes capable of degrading nylon oligomers. This was achieved in a mere nine days! The rapidity of this adaptation suggests a special mechanism for such adaptation, not something as haphazard as random mutations and selection."

Oh dear, it happened too fast. In actual fact, it was 9 days before colonies could grow at all on a simple nylon dimer, and three months before fast growing strains that could handle linear and cyclic dimers were isolated. This is typical of random mutation, a simple mutation allows the bacterial to just cope with the xenobiotic, allowing it a small selective advantage, and subsequent mutations improve on the initial weak activity. The time scale is not at all unusual for random mutation (if anything a bit slow).

"6. The researchers have not been able to ascertain any putative ancestral gene to the nylon-degrading genes. They represent a new gene family. This seems to rule out gene duplications as a source of the raw material for the new genes."

Not true, as before, the nylB group comes from a frameshift of an internally repetitious gene (so not surprisingly it is novel). NylA and NylC have not had homologous genes identified as of 2000, but then again a lot of bacterial sequencing has been done since, and as Don Batten states in a footnote, no Flavobacterium genome has yet been sequenced. Gene duplication is a major sources of new genes, but frame shifts, recombination and so on are all other sources of genes.

The whole article tries to show that the nylB (and other genes) cannot occur by random mutation, and must occur by directed mutation. They draw attention to B-cell hypermutation (which generates diversity in antibody genes) in vertebrates as an example of "directed" mutation. Unfortunately for them, hypermutation in B-cells is pure random shuffling with the occasional insertion, deletion and frame shift.

Generation of the nylon hydrolysing genes is standard "mutation followed by selection". The AiG article shows once again how poor their understanding of both biology and evolutionary theory is.


Negoro, S., Biodegradation of nylon oligomers (2000), Appl. Microbiol. Biotechnol. 54, 461-466.

Kato K, Ohtsuki K, Koda Y, Maekawa T, Yomo T, Negoro S, and Urabe I. (1995 Oct). A plasmid encoding enzymes for nylon oligomer degradation: nucleotide sequence and analysis of pOAD2. Microbiology, 141 (Pt 10), 2585-90.

Prijambada ID, Negoro S, Yomo T, and Urabe I. (1995 May). Emergence of nylon oligomer degradation enzymes in Pseudomonas aeruginosa PAO through experimental evolution. Appl Environ Microbiol, 61, 2020-2.

Yomo, T., Urabe, I. and Okada, H., (1992) No stop codons in the antisense strands of the genes for nylon oligomer degradation, Proceedings of the National Academy of Sciences USA 89, 3780-3784.

Kato K, Fujiyama K, Hatanaka HS, Priyambada ID, Negoro S, Urabe I, and Okada H. (1991 Aug 15). Amino acid alterations essential for increasing the catalytic activity of the nylon-oligomer-degradation enzyme of Flavobacterium sp. Eur J Biochem , 200, 165-9.

Ohno S. (1984 Apr). Birth of a unique enzyme from an alternative reading frame of the preexisted, internally repetitious coding sequence. Proc Natl Acad Sci U S A , 81, 2421-5

Ian Musgrave, Peta O'Donohue, Jack Francis, Michael James and Andrew Thomas Musgrave
Southern Sky Watch

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How to Have a Debate

Post of the Month Runner-Up: April 2004


Subject:    Re: Evolution is a proven fact it not ???
Date:       26 April 2004

"Roadrunner" wrote in message news:<YAajc.57286$>...

Let's stop beating around the bush and get right down to the core of the matter here, shall we? I'll begin.

If I thought that there was even a slight possibility that you could actually engage in a conversation I would spend several hours a day with you. But, unfortunately, in the past year you have shown that you can't engage in normal conversation. I don't know if that is because of a language barrier, a cultural barrier, or if you are just playing games but you have yet to have a single normal conversation despite your almost 2,000 posts to this group.

You may wish to claim that it is everyone else's fault that you are unable to have a normal conversation here but you know that isn't true otherwise no one else would be able to. When other people started to respond to your posts the same way that you have been responding to our posts you became frustrated so you can't claim that you don't realize what you are doing.

I am more than willing to share with you any source that I may use or derive information, a belief, or a position from. You are completely unwilling to share your sources other than Kent Hovind. I am willing to go into great detail on any subject but you are unwilling to do the same. Most people here are just as willing if not more so than I am to share with you but you are unwilling to share in return. This makes it impossible to have a normal discussion with you. Discussions are not a one way flow of information.

I would actually like to discuss several subjects with you but you are unwilling to discuss anything. Yes, you are all too eager to say that I am wrong or that most authority figures are wrong but that is as far as you have gone. You are not willing to discuss in detail why they are wrong.

You claim that I have a closed mind and that I need to investigate and study. Have you read any of my other posts in other threads? Have you read just how far I am willing to go in my search for the "truth"? I was a creationist and a Christian fundamentalist for almost 40 years of my life. Do you understand how much I have studied and how many things I have investigated "hands on" before I changed my beliefs? I doubt very much that you have taken the steps that I have taken to get where I am today.

You have read the things that we have said about you in this thread. They are not lies, they are not exaggerations. I take no pleasure in saying that you are unwilling to discuss anything. The things that we have said about you are based on your behavior here. All that you have to do is realize why we are saying these things and change your posting style so that we no longer have a reason to think this way. It's that simple.

Do you really want to have a discussion with anyone here?

If that is your goal do you realize why it has not happened to date?

Do you realize that you can't just tell people that they are wrong without good solid reasons based on evidence?

Do you realize that you can't just make up definitions for words that have a popular meaning in this newsgroup and then argue with your own definitions?

Do you realize that when someone points out that your source of material is wrong that you can't just wave your hands and continue using that source?

Do you realize that you will get nowhere here if you continue to publish lies told by Kent Hovind? Why should the people here respect someone who is repeating known lies?

If causing frustration is your only goal then you have accomplished it, otherwise in what way do you think that frustrating people will help either side?

Most people here are not playing a game. This is not a "win or lose" debating game. If that is what you are looking for then you would be better off watching TV game shows. If on the other hand you want to present information in a well thought out manner and if you are willing to discuss that information in detail, knowing that your information just may be wrong, and be willing to admit it then the beginnings of a discussion can take place. Otherwise you are just trolling for reactions to your nonsense of telling everyone that they are wrong.

I did not reach my level of understanding about biological evolution by watching TV shows or by reading pop culture books. I studied it in depth. I did not accept the age of the Earth by listening to a preacher or a slick public debater. I went out and studied different features and learned why those features take long lengths of time to form. I held those features in my hands and studied them in detail before I was willing to admit that the Earth is older than 6,000 years old. I held fast to my Christian fundamentalist beliefs but I was willing to learn and change those beliefs if proven wrong.

What are you willing to do to support your position?

How do you plan on teaching me that I am wrong when I was a creationist for longer than you have been alive? How do you plan on teaching me that I am wrong on subjects that I have spent years studying with hands on investigation? You will not teach me a thing by responding with "assumptions!" because I do not base my understanding on any assumption. You will not teach me anything by saying "investigate it!" because I have investigated it in depth. I am willing to bet that most people here have studied these things in a lot more depth than I have. How do you plan on teaching them that they are wrong? You will not be able to do it the way that you have been trying.

Until you present information in a reasonable manner and are willing to discuss that information to a depth acceptable by people who have already studied that information with hands on experience then you will get absolutely nowhere here and you will be wasting your time.

Do you understand why we are saying these things about you?
What are you willing to do in order to change the way we perceive you?
Do you understand why we don't get whatever point it is that you want to make?
Do you understand why biological evolution is not a religion?
Do you understand why we think the Earth is 4.5 billion years old?
Do you understand why we are frustrated with you?
Do you understand all of these things and everything else I have pointed out?

We do not want to insult you or make fun of you but when you continually act the way you have been you give people no other options. Ignoring you doesn't work because you insist on jumping into threads disrupting them with one line quips of nonsense like "assumptions!" and "evolution religion". Do you understand why I consider those posts of yours as nonsense? Am I wrong in classifying them as such?

Please be verbose in your response. Don't be afraid to type out full thoughts. Don't be afraid to get into detail. That's what is expected when you discuss an issue. I went into a little depth here. Can you match that depth in your response? Can you at least attempt to?

In fact I will give you a start. You claim that scientists continually test an object's age over and over again until the desired age is found. You claim to have witnessed this in a laboratory. But that is all that you will tell us. Most of us here find this a little hard to believe because this is not the way we have seen it done and we don't know any scientist that would actually do this. In fact if someone did do this at such a great expense we would consider it to be fraud on many different levels and would cry out against it wanting to see that scientist exposed. That's how outrageous your claim is and how serious it is.

Why do you refuse to give us any more information about it?
Why won't you tell us the actual dating method used?
What was the scientist dating?
Was there a scientific paper written on this and if so why won't you give us enough information so that we could find it and read it?
Do you know that it costs approximately $250 USD to carbon date an object? Did they date this object 10 times at a cost of $2,500? 100 times? 200 times?
Who paid for this fraudulent use?
How did the scientists hide the costs to date the object and avoid being terminated in disgrace from their position?
Did the lab own the approx. $3,000,000 dating equipment or did they send the sample out to another laboratory?
How old were you when you saw this done?
Why were you present during all of this?
Why did they allow you to be present during all this?
Why did the dates vary so much? Dates do not vary much if at all unless the object was contaminated. Then they would find a better sample to date, they wouldn't date the same sample again. You make it sound like they continually dated the same sample until they got the right date which would mean that not only do you not know how dating works but that you are not telling the truth about it.

These are just some of the questions that instantly came to mind when I read your claim. These questions need to be answered before we can believe that you are telling the truth and before we can discuss this further. That's how outrageous your claim is. Do you understand why we will not believe this claim without this information? Do you understand why your claim (whether true or not) is so outrageous?

If you do not understand why your claim is outrageous I will spell it out for you.

There was no reason for you to be present during the dating tests. None. The fact that you are claiming you were present raises some instant questions as to why you were present. One thought that instantly comes to mind is that the scientists were your parents. Other than that without more detailed information we can't understand any situation as to why you were present. This raises a big red flag.

Carbon dating costs about $250 a test. Multiple tests become instantly expensive. Someone has to pay for these tests. Most scientists couldn't afford them so that means that they are working for someone, a business or a school. Both track their expenditures closely and would never allow this to happen. If it did happen the scientists would be terminated. You claim that this is how all scientists date objects which obviously is not true. The fact that you claim this leads us to believe that this didn't really happen.

You will not receive different results if you date the same object. The only reason a new date would be needed is when the results obtained were obviously wrong meaning that the sample was contaminated. If dating an object that was found in the middle of a 16th century ruin comes back at 100 years old or 25,000 years old the sample has obviously been contaminated and a new sample would have to be found and tested. Despite the claims by anti-evolutionists dating methods are not inaccurate and will not give such varying results. This raises a big red flag. You obviously do not know how dating methods work and how expensive they are.

Since you have given us several reasons to doubt your knowledge pertaining to dating methods (and many other things) we find it incredibly difficult to believe that for some reason (which you refuse to give us) you were present during the lengthy dating procedures (hint: it takes more than 5 minutes to date an object) and that you were privy to the discussions by these fraudulent scientists as to the crime they were committing. And in a situation such as this most people would immediately report this fraudulent act to the next higher authority. You didn't. Instead you allowed it to happen and have done nothing about it other than claim the incident on a newsgroup claiming that this is how scientists work!

Now do you understand why we find this claim so incredibly difficult to believe and why we need so much more information about the incident before we could even begin discussing the matter with you? Many of your claims are just like this one making anything you claim suspect. But you want us to just swallow your stories hook, line, and sinker without questioning them. And then you have the nerve to claim that our minds are closed!

The ball is now in your court. Start explaining. Start giving us details. If you refuse to do this we have no other choice but to consider you not only ignorant of everything you speak of but worse, that you are a liar. We would rather not have this opinion about you because it is painful to think that a fellow human would actually behave like this but you give us no other option. Please prove us wrong.


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