Axe (2004) and the evolution of enzyme function

[Preface – the subject of protein evolution pops up on a regular basis in ID circles.  Recently, William Dembski mentioned the study alluded to in the title of this essay as an improved argument/piece of evidence for intelligent design.  Specifically, Dembski said:

“(2) The challenge for determining whether a biological structure exhibits CSI is to find one that’s simple enough on which the probability calculation can be convincingly performed but complex enough so that it does indeed exhibit CSI. The example in NFL ch. 5 doesn’t fit the bill. The example from Doug Axe in ch. 7 of THE DESIGN OF LIFE (www.thedesignoflife.net) is much stronger.”

“The example from Doug Axe in ch. 7 of THE DESIGN OF LIFE” would appear to be Axe’s 2004 paper in the Journal of Molecular Biology, the subject of my first ever essay on The Panda’s Thumb.  Since I have been a bit remiss in re-posting older essays here, I thought I would use this excuse to put this here.  It’s “published” without change, so as to maintain some sort of continuity.  As always, enjoy.]

Douglas Axe recently (well, sort of) published an article in the Journal of Molecular Biology entitled “Estimating the Prevalence of Protein Sequences Adopting Functional Enzyme Folds” (Axe, J Mol Biol 341, 1295-1315, 2004). In his discussion of the experimental observations, Dr. Axe mentions some numbers that are likely to generate much discussion amongst Intelligent Design advocates and critics. For example, Stephen Meyer (2004) cites Axe at a key point in the argument in his recent article advocating Intelligent Design, “The Origin of Biological Information and the Higher Taxonomic Categories,” much discussed in previous Panda’s Thumb threads (here).

“Axe (2004) has performed site directed mutagenesis experiments on a 150-residue protein-folding domain within a B-lactamase enzyme. His experimental method improves upon earlier mutagenesis techniques and corrects for several sources of possible estimation error inherent in them. On the basis of these experiments, Axe has estimated the ratio of (a) proteins of typical size (150 residues) that perform a specified function via any folded structure to (b) the whole set of possible amino acids sequences of that size. Based on his experiments, Axe has estimated his ratio to be 1 to 10^77. Thus, the probability of finding a functional protein among the possible amino acid sequences corresponding to a 150-residue protein is similarly 1 in 10^77.”

More recently, Dembski cited Axe in his Expert Witness Report for the Dover trial (see this).

“Recent research by Douglas Axe (see Appendix 3) provides such evidence in the form of a rigorous experimental assessment of the rarity of function-bearing protein sequences. By addressing this problem at the level of single protein molecules, this work provides an empirical basis for deeming functional proteins and systems of functional proteins to be unequivocally beyond Darwinian explanation.”

Given that this subject is often raised by ID proponents (such as this), and that the Biologic Institute (where Axe works) has made some news accounts, it seems appropriate to review Axe’s work. The purpose of this PT blog entry is to try and lay out the study cited above (Axe DD, J Mol Biol 341, 1295-1315, 2004) in a form that is accessible to most interested parties, and to discuss a larger context into which this work might be placed. Needless to say, the grand pronouncements being made by the ID camp are not warranted.

RNA and kitchen tools – Slicers, Dicers, and CRISPRs

RNA-based regulation is all the rage in biology today.  The more familiar mechanisms involve small RNAs such as microRNAs and silencing-associated RNAs.  The biogenesis and functioning of these RNAs involves enzymes and complexes that have been termed, among other things, Dicers and Slicers.  These subcellular kitchen utensils work by processing either the small RNA precursor or the base-paired target RNA.  This mode of regulation is most often associated with eukaryotes, and indeed homologous enzymes and mechanisms are not found in prokaryotes. However, systems with remarkable functional similarity may occur in bacteria.  A recent review by Sorek et al. brings one such example into focus.

One curious feature of bacterial genome is the occurrence of arrays of direct repeats in which the repeated units are separated by so-called spacers of unique sequence unrelated to the repeat units.  The sizes of the repeat units vary from bacteria to bacteria, ranging from between 24 to 47 bp.  Likewise, the spacer sizes vary from 26-72 bp.  These arrays are flanked by an apparent leader sequence, and yet again by arrays of protein-coding (CAS) genes, the number and composition of which vary considerably from bacteria to bacteria.  The general arrangement is shown in the following figure, which is part a of Figure 1 from Sorek et al. (shown beneath the fold):

The Murmansk Run

One of the lesser-known but compelling stories of the Second World War is that of the US Merchant Marine.  This service manned the convoys that plied the Atlantic and Pacific Oceans, and these sailors paid a heavy collective price in the war – I’ve read that some 1 in 26 sailors who served lost their lives in the conflict, a casualty rate greater than those of any of the branches of the armed services.  Perhaps the most harrowing of the routes plied by these convoys was the Murmansk Run – the route of convoys carrying supplies from the Western Allies to the Soviet Union, north around Norway to the Arctics port of Murmansk, Archangel, and Molotovsk.  This route was in easy reach of the U-Boat fleet and the surface navy of Germany.  Moreover, it was within easy striking distance of German bombers based in occupied Norway.  This meant that even heavily-escorted convoys were under perpetual threat.  Add to this the threat of the frigid Arctic waters, in which survival was a matter of minutes, and a picture of a harrowing and terrifying journey emerges.

One veteran of the Murmansk Run was my uncle, Clifton Bain.  Cliff passed away on Dec. 18, at the age of 92.  Cliff was a favorite brother of my mother’s, and he took great pleasure in calling me after the annual Kentucky-Florida football game (and more recently the basketball games) and gently needling me (readers should understand that Kentucky hasn’t beaten Florida in football since, oh, the days of the pharaohs).  I have happy memories of time spent with Cliff, but sadly not enough of these.  However, one stands out in my mind.  A few years ago, I took my family to Tampa to visit Cliff and his wife (Irma) for Christmas.  While we were there, Cliff took the time to tell us (and in particular Amy, who was in the middle of a school assignment having to do with chronicling the lives of our older citizenry) some stories of his younger days.  Among these was mention of his duty in the Merchant Marine and his experience on the Murmansk Run.  Incredibly enough, my mother, who was very close to Cliff, did not know that he had served there in WWII.  That he had never talked about these experiences with his sister struck me as unusual, and I realized after some reading that it was because of the horrors he likely endured.  Cliff was a strong and vibrant man, and it must have been an ordeal indeed to not share this with his sister (or others close to him).  This conveyed to me a sense of the terror of this duty that “book learning” never could.

I’ll miss Cliff, the annual phone call (not to mention that I’ll never have the opportunity to turn the tables and brag about that long-awaited Kentucky win over Florida in football), what he meant to my mother, and many other things.  But I’m thankful and blessed to have known him and had him as an uncle.

Strange things at promoters

A group of interesting papers popped up on ScienceExpress this past week.  These papers (by Core et al., Seila et al., He et al., and Preker et al.) all describe characterizations of unusual patterns of transcription in human cells.  The bottom line (well, one bottom line – there are lots of interesting data in these studies, and the nuances may take readers in slightly different directions) is that, for numerous promoters, transcription extends in both directions, not just in the one direction that is usually associated with productive (= leading to synthesis of a processed and translated mRNA) transcription.  Moreover, this bidirectional transcription is quite distinct from that associated productive transcription, in that it yields short and relatively unstable RNAs.  More elaboration follows below the fold.  As always, enjoy.

December 6, 1941

That’s not a typo. This essay is about the day before Pearl Harbor, a day that was of considerable importance in its own right (it was the culmination of what many deem the most important battle of WWII).

Dec. 6, 1941 was the date on which the Russian counteroffensive outside of Moscow began. This battle was important for many reasons, chief of which was that it was the first large-scale battlefield defeat suffered by the German Army in WWII. But beyond this (after all, larger wars are series of battlefield victories and defeats), it was the finishing touch of the growing portent for the German High Command that victory on the Eastern Front was at best a slim possibility.

Germany invaded Russia on June 22, 1941 and enjoyed considerable success in the early going. Success, to be sure, but at a price I do not think the German High Command expected. In terms of material and soldiers, the massive victories of the summer of 1941 were expensive, more than Germany could sustain over the course of years. But ultimate success seemed inevitable. By late summer, the numbers of killed, wounded, and captured Soviet troops were close to total estimates of manpower available to the Soviet Army in June of 1941. It probably was inconceivable to the German generals that the Soviet Union could continue to mount such dogged and costly defenses as they had done throughout the summer.

(The numbers are truly staggering. According to one source, by the end of August, Russia had lost more than 2.5 million soldiers . This was in one two-month campaign. By way of comparison, total US casualties for all of WWII, in both theatres, were about 1 million killed, wounded, or captured. The carnage on the Eastern Front is frightening to contemplate.)

But fight on the Soviet Army did. And with a ferocity (and cost in lives) that was unabated throughout the late summer and autumn. More defeats were suffered by Russia, much more territory lost, yet the battle raged. This had to concern the Germans, who by now were faced with an growing inhospitable climate as well as savage combat.

In spite of this, by late autumn, the German Army had slogged (and slugged) its way to the gates of Moscow, a prize whose fall was expected to be accompanied by a greater defeat of the remains of the Soviet Union. By early December, spent and ill-equipped though it was, the German Army was poised (or at least dug in, as much as could be done in the harsh winter conditions) to continue menacing Moscow, as they had been for the previous months.  December 6 changed this – the counteroffensive threw the Germans back from the gates of Moscow, and drove them back along a much larger front. Many incredibly bloody years awaited the combatants, years of brutal and horrifying actions on both parts.  But the die was largely cast on Dec. 6, that Germany could not defeat the Soviet Union.

(The scale of the overall Battle of Moscow, that lasted some seven months, was massive, larger than the more famous battles at Stalingrad and Kursk, as well as the Battle of the Bulge.  More than 7 million soldiers fought, and more than 2.5 million were killed, wounded, or captured.)

Off with their heads!

As noted in this earlier essay, the poly(A) tail collaborates with the 5′-end of the mRNA (the so-called cap) to promote both mRNA translation and stability.  Accordingly, decapping is a good hallmark for mRNA turnover.  In a recent issue of The Plant Cell, Jiao et al. describe an approach to study uncapped mRNAs on a global basis.  Briefly, these authors take advantage of the fact that an uncapped mRNA has a 5′ phosphate group, and thus can be a substrate for RNA ligase.  By attaching an RNA adapter to the uncapped mRNAs using this enzyme, and then purifying and amplifying DNA products derived from these, the authors were able to prepare probes for microarray studies.  Thus, they were able to assess uncapped mRNA abundance on a genome-wide basis.  As a test for this approach, they studied decapping genome-wide during the early stages of flowering using a mutant arrested for flower development at a specific stage, but carrying a chemically-inducible transgene that could trigger flowering by providing for some of the functionality missing in the mutant.  This group found that a sizable portion of mRNAs that could be detected on the microarrays (approaching 40%) were either “over-capped” or “under-capped”; that is to say, the relative abundances of uncapped mRNAs differed from the total levels of the corresponding transcripts.  They also found a number of transcripts (some 300 or so) whose capping status changed during flowering.  All told, as stated by the authors, this system should be useful for exploring regulated mRNA turnover, and for identifying correlations between mRNA sequence/structure and stability.

Thanksgiving in New York …

… or why entries have been slow in coming.